DENTAL  DEPARTMENT 


IN  MEMORIAL 

JOHN  ALBERT  MARSHALL,  D.D.S 


Putnam  's 
Science  Series 


1.  The  Study  of  Man.     By  A.  C.  HADDON. 

2.  The  Groundwork  of  Science.     By  ST.  GEORGE  MIVART. 

3.  Rivers  of  North  America.    By  ISRAEL  C.  RUSSELL. 

4.  Earth  Sculpture,  or  ;  The  Origin  of  Land  Forms.    By  JAMES 

GEIKIE. 

5.  Volcanoes ;  Their  Structure  and  Significance.    By  T.  G. 

BONNEY. 

6.  Bacteria.     By  GEORGE  NEWMAN. 

7.  A  Book  of  Whales.     By  F.  E.  BEDDARD. 

8.  Comparative  Physiology  of  the  Brain,  etc.  By  JACQUES  LOEB. 

9.  The  Stars.     By  SIMON  NEWCOMB. 

10.  The  Basis  of  Social  Relations.     By  DANIEL  G.  BRINTON. 

n.  Experiments  on  Animals.     By  STEPHEN  PAGET. 

12.  Infection  and  Immunity.     By  GEORGE  M.  STERNBERG. 

13.  Fatigue.     By  A.  Mosso. 

14.  Earthquakes.     By  CLARENCE  E.  BUTTON. 

15.  The  Nature  of  Man.     By  ELIE  METCHNIKOFF. 

16.  Nervous  and  Mental  Hygiene  in  Health  and  Disease.     By 

AUGUST  FOREL. 

17.  The  Prolongation  of  Life.    By  ELIE  METCHNIKOFF. 

18.  The  Solar  System.     By  CHARLES  LAKE  POOR. 

19.  Heredity.     By  J.  ARTHUR  THOMPSON,  M.A. 
ao.  Climate.     By  ROBERT  DECOURCY  WARD. 

ai.  Age,  Growth,  and  Death.    By  CHARLES  S.  MINOT. 

as.  The  Interpretation  of  Nature.     By  C.  LLOYD  MORGAN. 

33.  Mosquito  Life.    By  EVELYN  GROESBEECK  MITCHELL. 

34.  Thinking,  Feeling,  Doing.     By  E.  W.  SCRIPTURE. 

35.  The  World's  Gold.     By  L.  DE  LAUNAY= 

36.  The  Interpretation  of  Radium.     By  F.  SCDDY. 

27.  Criminal  Man.     By  CESARE  LOMBROSO. 

28.  The  Origin  of  Life.     By  H.  CHARLTON  BASTIAN. 

29.  The  Bacillus  of  Long  Life.        By  LOUDON  M.  DOUGLAS.  ' 

30.  Social  Evil.      By  R.  A.  SELIGMAN. 

31.  Microbes  and  Toxins.      By  E.  BURNET. 

For  list  of  works  in  preparation  see  end  of  this  volume 


I 


PASTEUR 

When  a  pupil  at  the  Ecole  Normale,  1843-46 
Pencil  drawing  by  Lebayle  from  a  daguerreotype  in  the  possession  of  M.  Vallery-Radot 


MICROBES  y  TOXINS 


< 


By  Dr.  ETIENNE    BURNET 

OF   THE    PASTEUR    INSTITUTE   OF    PARIS 
\YITH  A  PREFACE  BY 

ELIE   METCHNIKOFF 


Translated  from  the  French 

BY 

Dr.  CHARLES   BROQUET  and  W.  M.  SCOTT,  M.D 


ILLUSTRATED 


G.  P.  PUTNAM'S  SONS 

NEW  YORK  AND  LONDON 
Imicfeerbocfeer   press 
1912 


TO 

DR.  EMILE   ROUX 

Director  of  the  Institut  Pasteur 


v',V 


Photo] 


ILLIE  METCHNIKOFF 


\Henri  Manuel,  Paris. 


INTRODUCTION 


THE  publication  in  the  Bibliotheque  de  Philosophic  Scientifique 
of  a  volume  dealing  with  micro-organisms  was  entirely  indicated, 
for  microbiology  is  taking  every  day  a  larger  and  larger  place 
in  the  realm  of  knowledge  and  philosophy.  Although  dis- 
covered more  than  two  hundred  years  ago,  microbes  were  long 
neglected,  and  it  was  only  during  the  second  half  of  last 
century  that  their  true  role  was  ascertained. 

Ehrenberg  in  the  middle  of  the  nineteenth  century  had 
already  perceived  the  importance  of  microscopic  organisrrs  in 
the  evolution  and  life  of  our  planet.  The  discovery  of  the 
fossil  remains  of  Diatoms  and  Foraminifera  led  him  to  appre- 
ciate the  great  part  these  minute  creatures  have  played  in  the 
building  up  of  the  earth's  crust. 

There  were  not  lacking  men  of  science  disposed  to  attribute 
to  micro-organisms  an  important  action  in  the  phenomena  of 
fermentations  and  of  disease,  but  it  was  only  after  the  labours 
of  Pasteur  that  this  truth  was  definitely  established  and  became 
part  of  our  common  heritage  of  knowledge. 

Relations  had  long  been  perceived  between  the  animal 
kingdom  and  the  vegetable,  animals  furnishing  carbonic  acid 
and  nourishment  to  plants,  while  these,  on  their  side,  nourish 
the  animals  with  their  organic  matter  and  provide  them  with 
oxygen.  Later  it  was  recognised  that  between  these  two 

kingdoms  there  lies  the  domain  of  the  microbes.     It  is  the 

u 


x  INTRODUCTION 

microbe  which  transforms  the  animal  material  supplied  by 
dead  bodies  and  dejecta  into  simpler  substances,  nitrates,  and 
salts  of  ammonia,  capable  of  assimilation  by  those  plants  which 
supply  us  with  food.  Further,  it  is  the  microbe  which  renders 
pleasant  to  the  taste  certain  animal  and  vegetable  food- 
products,  as,  for  example,  the  juice  of  the  grape,  the  extract 
of  malt,  cabbages,  apples,  and  milk,  transforming  these 
respectively  into  wine,  beer,  sauerkraut,  cider,  kephir,  various 
kinds  of  cheese,  etc. 

Thanks  to  Pasteur  the  activity  of  micro-organisms  was 
established  in  every  case  of  putrefaction  and  fermentation ; 
and,  with  this  fact  to  start  from,  it  became  more  easy  to  tackle 
the  problem  of  infectious  diseases. 

Putrefaction  and  suppuration  have  been  recognised  for  cen- 
turies as  being  phenomena  of  the  same  order.  Decomposing 
pus,  faecal  matter  smelling  of  putrefaction,  urine  issuing  from 
the  bladder  in  a  state  of  decomposition,  all  indicated  that  an 
illness,  a  state  of  suffering,  was  equivalent  to  an  infection  of  the 
body. 

Although  certain  microbes,  such  as  the  bacteridium  of  anthrax, 
had  been  observed  before  Pasteur's  great  discoveries,  it  was  only 
as  a  consequence  of  these  discoveries  that  the  fundamental  role 
of  microscopic  organisms  in  disease  was  understood.  The 
labours  of  Lister  in  surgery,  and  of  Davaine  and  Koch  on  the 
"  black  blood  of  anthrax  "  first  authorised  the  application  of 
Pasteur's  doctrine  to  surgery  and  medicine. 

Pasteur  himself  with  his  pleiad  of  disciples  was  in  the  midst 
of  this  activity,  an  activity  which  in  a  very  few  years  revolu- 
tionised medical  science  and  endowed  medicine  with  more 
than  one  preventive  vaccine  such  as  those  against  anthrax  and 
rabies. 

The  Pasteur  school  in  France  and  the  school  of  Koch  in 
Germany  have  succeeded  in  elucidating  many  medical  problems 
of  the  highest  importance  and  have  drawn  valuable  practical 
conclusions  from  these. 

Thanks  to  all  this  work,  work  which  has  increased  during 
these  last  years  in  extraordinary  fashion,  a  universe  of  micro- 


INTRODUCTION  xi 

organisms,  beneficent  and  mischievous,  has  been  revealed  to 
humanity ;  and  it  is  this  new  knowledge  which  has  so  largely 
contributed  to  the  diminution  in  disease  and  death  at  the 
present  day  and  which  holds  out  to  man  the  hope  of  a  more 
happy  future. 

The  micro-organisms  inhabiting  our  bodies  have  set  going 
there  a  poison  factory,  which  cuts  short  our  existence,  and  by 
secreting  poisons  which  penetrate  all  our  tissues,  injures  our 
most  precious  organs,  our  arteries,  brain,  liver,  and  kidneys. 

Man  balked  of  his  full  term  of  life  feels  himself  unhappy  and 
is  ready  to  accept  any  solution  to  the  problem  of  gaining 
happiness.  And  the  progress  of  microbiology  leads  us  to  hope 
that  this  science  will  one  day  liberate  man  from  his  fear  of  the 
grave  and  permit  him  to  attain  the  true  object,  the  true 
conclusion  of  life. 

It  is  time  for  bacteriological  science  to  leave  the  laboratory 
and  the  lecture  theatre,  and  to  take  its  place  before  the  great 
public,  in  order  that  its  benefits  may  receive  the  widest  and 
readiest  application. 

It  was  very  natural  for  the  creator  of  this  "  Library  of 
Scientific  Philosophy  "  to  apply  to  the  Pasteur  Institute  for  an 
account  of  the  actual  position  of  science  with  regard  to  microbes 
and  toxins.  Not  only  was  the  movement  started  from  Pasteur's 
laboratory  and  continued  in  the  Institute  bearing  his  name, 
and  still  sheltering  one  of  his  most  illustrious  collaborators  in 
the  person  of  Dr.  Roux,  but  it  is  in  this  Institute  that  every 
branch  of  microbiology  is  undergoing  active  study.  To  take 
colloids  and  the  physico-chemical  laws  which  govern  their 
activity,  we  have  at  the  Institut  Pasteur  studies  on  ferments 
and  fermentations  as  well  as  on  the  chemical  processes  which 
lie  at  the  root  of  life  and  of  recovery  from  disease.  In  this 
Institute  also  there  are  zealous  workers  in  the  field  of  infective 
microbes  and  the  means  of  combating  them. 

Several  laboratories  are  specially  set  apart  for  researches  on 
tropical  diseases,  and  finally  the  Pasteur  Hospital  has  been 
created  for  patients  suffering  from  all  sorts  of  infectious  maladies. 

If  Pasteur  were  to  see  his  Institute  again,  he  would  be 


xii  INTRODUCTION 

astonished  at  the  great  changes  which  have  taken  piace  in  it 
and  in  the  science  of  microbiology  in  general.  It  would  take 
him  some  time  to  overtake  and  realise  the  progress  attained. 
And  yet  in  spite  of  all  that  has  been  done  there  remains  still 
much  work  for  the  future.  Many  scourges  still  await  a  remedy. 
In  the  case  of  tuberculosis,  although  extraordinary  advances 
have  been  made  in  its  study,  the  final  solution  of  the  problem 
is  still  reserved  for  the  future.  The  great  question  of  cancer, 
so  important  and  so  difficult,  has  been  hardly  more  than  opened. 
There  remain  to  be  discovered  the  microbes  of  many  diseases, 
e.g.,  hydrophobia,  scarlatina,  and  measles,  which  are  perhaps 
filtrable  micro-organisms,  invisible  with  the  best  microscopes. 

The  field  of  infectious  diseases  is  extending  wider  and  wider 
with  the  progress  of  microbiology.  We  find  that  certain  dis- 
eases thought  to  be  diseases  of  metabolism  are  beginning  to  be 
classed  in  this  group.  Arterio-sclerosis,  an  affection  so  wide- 
spread and  so  apt  to  cut  short  our  existence,  results  from  the 
activity  of  our  intestinal  flora.  Perhaps  before  long  it  will  be 
possible  to  explain  diabetes,  gout,  and  rheumatism  by  the 
injurious  activity  of  some  variety  of  microbe. 

Even  in  those  problems  of  hygiene  which  affect  society  in 
general  microbiology  is  taking  the  predominant  role.  The 
grand  problem  of  a  rational  food  supply,  which  used  to  be 
thought  capable  of  solution  by  inventions  of  chemistry  and 
physics,  will  necessarily  have  to  be  studied  by  microbiological 
methods,  in  view  of  the  fact  that  the  intestinal  bacteria  play 
one  of  the  most  important  parts  in  everything  that  concerns 
nutrition.  It  is  not  sufficient  to  state  the  nutritive  value  of  a 
food  in  terms  of  the  calories  which  it  contains  ;  it  will  still  be 
necessary  to  define  precisely  its  relations  to  the  intestinal  flora 
from  the  point  of  view  of  the  production  of  microbic  poisons. 

M.  Gustave  le  Bon  asked  me  to  put  together  in  a  little 
volume  for  his  Library  a  summary  of  what  is  known  about 
microbes  and  toxins.  I  advised  him  to  apply  rather  to  one  of 
my  young  colleagues,  and  I  indicated  in  particular  Dr.  Burnet. 
I  am  happy  to  find  that  I  could  not  have  chosen  better.  In 
spite  of  the  great  difficulties  there  are  in  attempting  to  describe 


INTRODUCTION  xiii 

within  a  limited  space  the  result  of  the  innumerable  labours 
which  have  accumulated  on  microbes  and  their  poisons  and 
which  have  been  pursued  in  the  most  varied  directions, 
M.  Burnet  has  succeeded  in  accomplishing  the  task  in  a 
remarkable  fashion.  I  am  sure  that  those  who  read  will  share 
my  opinion,  and  I  wish  this  book  the  most  widespread 
popularity. 

ELIE    METCHNIKOFF. 
PARIS,  1911. 


CONTENTS 


CHAPTER   I 

PACK 
THE   GENERAL   FUNCTIONS  OF    MICROBES — THE  TRANSFORMATION 

CYCLES  OF  CARBON   AND  NITROGEN  I 


CHAPTER   II 

MICROBES     IN    THE    HUMAN     BODY— LIFE     WITHOUT     MICROBES  — 

THE    INTESTINAL   FLORA 25 


CHAPTER  III 

FORM  AND   STRUCTURE    OF   MICROBES $O 

CHAPTER  IV 

PHYSIOLOGY   OF  THE   MICROBES 72 

CHAPTER  V 

PATHOGENIC   MICROBES— INFECTION IO7 

CHAPTER  VI 

INFLAMMATION    AND    PHAGOCYTOSIS 130 

CHAPTER  VII 

THE   PATHOGENIC   PROTOZOA:    FILTER-PASSING   VIRUSES  ...      14! 

xv 


xvi  CONTENTS 

CHAPTER  VIII 

PAGE 

THE  TOXINS 157 

CHAPTER  IX 

TUBERCULIN   AND   MALLEI N— ANIMAL  TOXINS— VENOMS  .          -173 

CHAPTER  X 

IMMUNITY l8; 

CHAPTER  XI 

IMMUNITY 210 

CHAPTER  XII 

ANAPHYLAXIS 233 

CHAPTER  XIII 

APPLICATIONS  OF  BACTERIOLOGY 250 

CHAPTER  XIV 

VACCINES  AND  SERA 265 

CHAPTER  XV 

CHEMICAL  REMEDIES 285 


MICROBES    AND    TOXINS 


CHAPTER  I 

THE  GENERAL  FUNCTIONS  OF  MICROBES — THE  TRANSFORMATION 
CYCLES  OF  CARBON  AND  NITROGEN 

The  circulation  of  matter  ;  anabolism  and  catabolism — Views  of  Lavoisier — 
The  transformation  cycles  of  carbon  and  nitrogen — The  role  of  micro- 
organisms :  I.  Maturation  of  the  soil  and  formation  of  arable  land. 
II.  Fermentation  of  vegetable  matter — Decomposition  of  starch  and 
manure — Hypotheses  on  the  formation  of  coal.  III.  Putrefaction  of 
albuminous  materials.  IV.  Nitrification  and  denitrification :  in  agricul- 
ture ;  in  the  biological  purification  of  sewage.  V.  Fixation  of  atmos- 
pheric nitrogen  in  the  soil :  by  bacteria  alone ;  by  bacteria  in 
association  with  algae  ;  by  the  nodule  bacteria  of  Leguminosse — 
Some  ideas  about  the  useful  micro-organisms :  Fermentations  in  con- 
nection with  food  production  and  in  various  industries. 

ALMOST  all  living  matter  is  made  up  of  water,  that  is  to  say, 
of  oxygen  and  hydrogen,  and  of  compounds  of  carbon  and 
nitrogen.  Other  elements  may  enter  into  the  tissues  of 
animals  and  vegetables,  for  example,  sulphur,  iron,  arsenic, 
boron;  but  by  following  the  circulation  of  carbon  and 
nitrogen,  it  is  possible  to  have  a  general  view  of  the  movements 
of  exchange  between  matter  and  life. 

The  living  creature  restores  to  nature  what  it  has  absorbed, 
and  eventually  its  own  body,  by  its  excretions  during  life  and  by 
its  decomposition  after  death.  The  elements  set  free  are 
recombined  into  organic  bodies,  and  these  exchanges  and  this 
circulation  form  the  essence  of  life.  It  is  an  abuse  of  our 
subjective  attitude  to  consider  as  two  opposites  life  and  death. 
At  most,  life  is  the  opposite  of  inertia.  Death  is  a  special  kind 
of  accident,  life  being  the  all-embracing  phenomenon.  The 
first  of  the  workers  in  the  great  cycle  of  life  are  the  microbes, 


2  MICROBES   AND  TOXINS 

and  the  decompositions  and  re-combinations  of  living  matter 
depend  entirely  upon  them. 

Life  without  microbes  is  not  conceivable  at  the  present 
day.  That  does  not  mean  that  they  were  the  first  living  beings 
to  appear  on  the  surface  of  the  earth.  It  is  the  more  difficult 
to  get  an  idea  of  their  origin,  since  in  all  probability  they  have 
undergone  evolution,  and  have  not  always  had  the  appearance 
they  have  at  present.  It  is  possible  that  under  forms  that  we 
can  hardly  guess  at,  life  appeared  long  before  the  existence  of 
microbial  forms ;  but  microbes  have  been  the  chief  agents  in 
the  spread  and  extension  of  life  throughout  the  world. 

In  the  world  of  to-day,  the  building  up  and  breaking  down 
of  organic  substances  are  functions  of  cells,  conducted  by  an 
infinite  number  of  diastases.  These  cells  include  the  cells 
built  up  into  animal  and  vegetable  tissues,  and  the  isolated 
cells,  the  microbes.  It  would  not  be  right  to  contrast  too 
much  the  microbial  cells  with  the  cells  of  tissues,  merely 
because  the  former  are  separate  individuals,  whereas  the  latter 
are  arranged  in  systems.  In  nature,  micro-organisms  can 
exist  as  solitary  individuals,  but  are  seldom  actively  at  work  in 
this  condition.  For  example,  in  the  soil,  the  bacteria  which 
fix  nitrogen  act  rather  like  a  wide-spread  tissue.  The  active 
agents  are  in  any  case  invariably  cells. 

To  describe  in  a  few  words  the  cycle  which  organic  matter 
follows,  one  cannot  do  better  than  quote  the  celebrated  page 
of  Lavoisier,  "  Plants  draw  from  the  circumambient  air,  from 
water  and  in  general  from  the  mineral  kingdom,  the  substances 
necessary  for  their  own  organisation.  Animals  feed  either  on 
plants  or  on  other  animals,  which  have  themselves  fed  on 
plants,  so  that  eventually  the  matter  building  them  up  is 
always  derived  either  from  the  air  or  from  the  mineral 
kingdom. 

"Finally,  fermentation,  putrefaction  and  combustion  are 
continually  restoring  to  the  atmosphere  and  to  the  mineral 
kingdom  the  elements  which  plants  and  animals  have 
borrowed. 

"By    what      recesses    does   nature    effect   this    wonderful 


GENERAL  FUNCTIONS   OF  MICROBES        3 

circulation  between  the  two  kingdoms?  How  does  nature 
succeed  in  producing  substances  which  are  combustible, 
putrescible  and  capable  of  fermentation  from  combinations 
which  have  none  of  these  properties  ?  Here  are  impenetrable 
mysteries.  One  may  perceive,  however,  that  since  combustion 
and  putrefaction  are  the  means  which  nature  employs  to  return 
to  the  mineral  kingdom  what  has  been  drawn  from  it  in  the 
building  up  of  plants  and  animals,  the  latter  process  must  be 
the  converse  of  the  former." 

Green  plants  get  their  carbon  from  the  carbonic  acid  of  the 
air.  In  virtue  of  their  chlorophyll  activity  they  build  up  this 
carbon  into  starch,  cellulose,  sugars  and  fats.  This 
carbohydrate  synthesis  then  represents  the  accumulation  of 
energy.  A  green  plant  kept  in  the  dark  burns  up  its 
hydrocarbons,  returning  them  to  the  atmosphere.  A  dead 
plant  returns  its  hydrocarbons  after  a  series  of  decompositions, 
in  the  form  of  carbonic  acid  and  water.  A  plant  which  has 
been  eaten  by  an  animal  supplies  the  animal  with  glycogen  and 
fats  which  are  consumed  in  the  course  of  muscular  work  and 
respiration.  Dying  animals  like  dead  plants  return  their 
hydrocarbons  to  nature.  Plants  without  chlorophyll  and  all 
animals  spend  and  dissipate  the  energy  accumulated  by  the 
chlorophyllous  plants,  energy  derived  entirely  from  the  rays 
of  the  sun. 

Nitrogen  exists  in  the  atmosphere,  from  which  it  enters  the  soil. 
It  exists  in  the  dejecta  of  living  animals  and  in  the  bodies  of 
plants  and  animals  rotting  on  the  surface  of  the  earth.  It 
accumulates  there  in  the  mould  or  humus  :  plants  take  up  the 
nitrogenous  matter  of  the  soil  in  the  form  of  salts  of  ammonia 
and  nitrates,  and  build  up  from  them  the  vegetable  proteins. 
Animals  which  eat  plants  produce  from  these  the  animal 
proteins.  Animals  and  plants  in  decomposition,  and  the 
excretions  of  animals  in  general,  return  their  nitrogen  to 
fertilize  the  soil. 

Animals  are  in  a  certain  sense  parasites  of  plants,  since  they 
are  unable  to  build  up  by  themselves,  starting  from  mineral 
elements,  their  hydrocarbons  and  proteins.  Even  in  plants 

B    2 


4  MICROBES    AND   TOXINS 

the  formation  of  the  protein  compounds  is  dependent  on  the 
chlorophyll  activity;  without  this  it  would  be  impossible 
for  them  to  build  up  the  carbohydrates,  and  these  are 
indispensable  in  the  employment  and  elaboration  of  the 
nitrogenous  elements  of  the  soil.  The  chlorophyll  activity 
is  itself  dependent  on  the  sunlight,  so  that  life  is  one  great 
paean  to  the  sun. 

In  the  transformation  of  carbon  and  nitrogen  all  the 
operations  are  not  carried  out  by  microbes,  for  these  latter  do 
not  count  at  all  in  chlorophyll  assimilation  nor  in  animal 
digestion.  But  it  is  the  bacteria  which  keep  up  the  supply  of 
organic  matter  which  forms  the  source  of  animal  and 
vegetable  life ;  it  is  the  bacteria  which  restore  to  circulation 
those  elements  which  were  for  a  moment  arrested  in  the  bodies 
of  animals  and  plants :  they  restore  to  life  matter  which  had 
ceased  to  live.  It  is  they  also  which  prepare  the  soil  for 
vegetation  and  cultivation  :  they  accumulate  in  the  soil  the 
nitrogen  which  we  are  to  eat  in  the  form  of  cereals. 

It  is  they  which  carry  on  the  impenetrable  mystery  referred 
to  by  Lavoisier. 

They  prepare  the  primary  material  of  life.  Their  activity 
is  as  universal  as  that  of  water,  as  that  of  light. 

The  discoveries  of  Pasteur  have  not  only  revolutionised 
medicine,  they  have  rilled  with  new  life  the  science  and 
practice  of  agriculture  and  husbandry. 

I. — Maturation  of  the  soil  and  formation  of  arable  land. 

It  was  not  the  bacteria  which  shattered,  splintered  and 
powdered  the  rocks  which  formed  the  first  crust  of  the  cooling 
globe.  But  as  soon  as  in  this  chaos  there  appeared  water  and 
alkaline  phosphates,  algae  and  bacteria  could  establish 
themselves  and  with  the  help  of  carbonic  acid  continue  the 
first  formation  of  soil. 

In  those  rocks  known  as  "  rotten  rock,"  Miintz  found 
nitrifying  microbes  :  the  nitrifying  ferment  has  also  attacked 
the  Faulhorn,  a  mountain  of  the  Bernese  Oberland  near 


GENERAL  FUNCTIONS   OF  MICROBES        5 

Grindelwald,  which  is  composed  of  a  calcareous  schist,  black, 
friable,  and  crumbling  down.  There  is  therefore  a  sort  of 
putrefaction  of  rocks  (alkalis  and  alkaline  earths).  According 
to  Fausto  Sestini  the  carbonic  acid  produced  in  the  respira- 
tion of  plant  roots  hastens  the  breaking  down  of  feldspars. 

Mosses,  lichens,  algae  and  bacteria  prepare  the  way  for  the 
highest  plants.  The  more  there  is  growth  of  plants,  the  more 
there  is  decomposition.  In  this  way,  soil  or  humus  has  little 
by  little  spread  over  all  the  earth. 

II. — Fermentation  of  Carbohydrates. 

These  fermentations  restore  to  circulation  the  hydrocarbons 
of  animals  and  plants. 

For  the  decomposition  of  sugars,  starches,  fats,  glucosides, 
and  celluloses,  several  series  of  fermentations  are  necessary, 
each  supplementing  another  in  the  work.  The  anaerobic 
organisms  break  down  the  large  organic  molecules,  whereas 
the  aerobes  carry  out,  in  particular,  oxidations.  When  the 
bacteria  are  insufficient,  the  moulds  continue  the  work. 
Finally,  nothing  is  left  but  carbonic  acid  and  water.  All  the 
operations  which  in  our  laboratory  analyses  we  distinguish  and 
conceive  in  terms  of  abstract  formulae,  are  combined  and 
intermingled  in  nature,  each  succeeding  and  limiting  the  other. 
The  yeasts  transform  sugars  into  carbonic  acid  and  alcohol. 
Alcohol  attacked  by  the  acetic  ferments  is  turned  into  acetic 
acid.  Finally,  the  Bacterium  aceti  can  split  the  acetic  acid 
of  vinegar  into  carbonic  acid  and  water.  The  Mycoderma  vim 
destroys  and  oxidises  both  alcohol  and  acetic  acid,  producing 
again  carbonic  acid  and  water.  The  sugar  contained  in 
Raulin's  fluid  (v .  infra},  for  example,  is  attacked  by  Aspergillus 
niger,  the  products  including  oxalic  acid.  Various  moulds  and 
bacteria  turn  starch  into  sugar ;  it  is  the  moulds  in  particular 
which  complete  the  oxidations,  producing  carbonic  acid  and 
water. 

Milk  left  to  itself  ferments,  i.e.,  produces  lactic  acid,  which, 
meeting  with  an  alkaline  carbonate,  furnishes  calcium  lactate. 


6  MICROBES   AND  TOXINS 

Pasteur  discovered  the  transformation  of  the  lactate  into  the 
butyrate  by  the  butyric  vibrio ;  the  butyrate  can  be  completely 
consumed  in  its  turn  by  the  moulds. 

The  glucosides  (compounds  of  sugar  with  an  organic  body, 
an  alcohol  or  phenol)  are  decomposed  into  their  two  elements. 
A  diastase,  tannase,  secreted  by  the  Aspergillus,  splits  tannin 
into  two  molecules  of  gallic  acid,  from  which  other  moulds 
produce  again  carbonic  acid  and  water. 

Just  as  there  is  not  one  starch  but  several,  so  there  are 
several  celluloses,  which  resemble  starches  but  are  more 
stable :  (CgHjQOg)^.  They  form  the  walls  of  vegetable  cells, 
and  make  up  one-third  of  the  weight  of  the  straw,  which  is 
the  principal  component  of  farmyard  manure.  If  the  celluloses 


FIG.  I. — The  microbe  which  ferments  cellulose,  described  by  Omeliansky : 
bacilli  with  spores. 

were  not  decomposed  and  restored  to  circulation,  the  earth 
would  soon  be  cumbered  with  useless  refuse  material.  But  from 
the  beginning  moulds  establish  themselves  on  the  outer  skin  of 
the  living  plant ;  when  it  dies  they  invade  its  tissues,  attacking 
first  the  sugar  and  then  the  cellulose,  the  latter  being  hydrolysed, 
transformed  into  sugars  and  consumed.  The  B.  Amylobacter 
of  Van  Tieghem,  an  anaerobe,  produces  from  cellulose 
hydrogen,  carbonic  acid  and  butyric  acid.  Omeliansky  has 
demonstrated  two  methods  of  anaerobic  fermentation  in 
cellulose,  one  with  production  of  hydrogen,  the  other  with 
production  of  methane.  Those  ferments  which  in  a  tube  in 
the  laboratory  decompose  the  cellulose  of  Berzelius'  paper, 
act  in  precisely  the  same  way  in  manure  heaps.  The  aerobic 
fermentation  of  cellulose  is  carried  on  by  moulds,  by  fungi 


GENERAL  FUNCTIONS   OF  MICROBES        7 

more  highly  organised,  and  by  the  nitrifying  and  denitrifying 
bacteria. 

Pectose  is  a  hydrocarbon  associated  with  cellulose  in  the 
membranes  and  interstices  of  plant  cells;  the  rust  which 
attacks  hemp  and  linen  is  a  fermentation  of  the  pectose, 
transforming  it  into  pectic  acid,  then  to  sugar,  by  the  B. 
amylobacter  and  by  the  Granulobacter  of  Fribes  and  Winograd- 
sky.  In  the  manure  heap,  aerobic  fermentation  proceeds  at 
the  surface  and  raises  the  temperature  up  to  nearly  80°  C.  In 
the  depths  of  the  heap  the  temperature  is  low,  and  there  the 
anaerobic  ferments  attack  the  cellulose.  The  bubbles  which 
rise  and  burst  on  the  surfaces  of  ponds  are  signs  of  the 
anaerobic  fermentation  which  is  decomposing  organic  debris  at 
the  bottom.  Manure  kept  in  a  latticed  box  with  free  access  of 
air  heats  up  without  there  being  destruction  of  the  cellulose ; 
loss  of  nitrogen  takes  place.  Manure  heaped  in  a  closed '  box 
or  kept  corked  in  a  large  carboy,  liberates  methane  from  the 
decomposition  of  the  cellulose.  It  is  possible  to  collect  the 
gas  and  by  means  of  an  exit  tube  to  make  it  furnish  a  light. 

It  was  a  natural  step  to  attribute  to  the  ferments  of 
cellulose  the  formation  of  peat,  lignite  and  coal.  Microbes 
certainly  are  at  work  in  the  decomposition  of  vegetable  matter 
in  the  peat-bogs,  and  coal  is  supposed  to  be  the  product  of  more 
complete  fermentations,  two  varieties  at  least  of  bacteria  being 
in  activity  in  succession;  the  first  dissolves  the  central 
membranes  of  the  cell  walls,  the  others  attacking  the  cellulose, 
more  or  less  pure,  which  constitutes  the  thick  parts  of  the  wall. 
"  The  bacterial  activity,"  says  the  most  convinced  defender  of 
this  theory,  "produced  a  de-hydrogenation  and  de-oxidation, 
the  final  result  being  the  production  of  carbon.  We  do  not 
know  if  the  final  limit  of  this  process  has  ever  been  reached, 
but  figures  show  that  the  more  geologically  ancient  the  fuel,  the 
more  carbon  it  contains  "  (Bernard  Renault).  Fuel  of  less 
age,  lignite,  and  peat,  for  example,  contains  besides  bacteria, 
amoebae  and  infusoria.  In  the  lignites,  the  cannel  coals,  and  the 
boghead  coals,  we  find  fungi  and  algae,  which  do  not  occur  in 
coal  proper.  Bacteria  alone  exist  in  all  the  fuels ;  they  are 


8  MICROBES   AND   TOXINS 

much  less  altered  than  the  structures  which  surround  them  : 
therefore  it  is  supposed  that  they  must  have  survived  tissues 
which  they  destroyed. 

The  presence  of  bacteria  in  coals  derived  from  organic 
matter  should  not  astonish  us.  But  it  appears  difficult  to 
believe  that  bacteria  have  been  the  only  agents  in  the  formation 
of  coal.  The  study  of  their  form  is  extremely  difficult ;  fine 
particles  of  iron  pyrites  and  little  crystals  often  imitate  bacterial 
forms  in  the  thin  sections  of  coal,  so  difficult  to  prepare  and 
examine  by  transmitted  light  under  the  microscope.  On  the 
other  hand  the  fermentation  of  cellulose  does  not  explain  the 
frequent  impregnation  of  the  debris  by  a  blackish,  bituminous 
material,  nor  perhaps  does  it  really  explain  the  enrichment  in 
carbon  of  the  deposits ;  further,  there  have  been  produced  from 
the  fermentation  of  fatty  bodies,  resins  and  similar  volatile 
substances,  phenol  bodies  like  those  found  in  coal  (E.  Duclaux). 
There  is  nothing  surprising  in  the  presence  of  algae  in  the 
Bogheads^  for  these  are  precisely  the  coals  derived  from  algae 
and  often  named  algal  coals.  Their  formation  was  due  to 
luxuriant  algae  growths  rapidly  developed  on  the  surface  of 
stagnant  pools,  which  then  sank  to  the  bottom  carried  down 
by  a  coagulum  in  the  peaty  water ;  this  mass  contained 
bituminous  substanceswhich  must  have  come  from  elsewhere,  for 
there  is  no  indication  that  they  took  their  origin  on  the  spot  by 
an  alteration  of  the  algae.  It  is  these  bitumens  which  have 
produced  the  carbon  enrichment  of  the  mass,  so  that  on  the 
whole  instead  of  destruction  a  preservation  process  has  taken 
place.  In  other  cases,  instead  of  this  enormous  growth  of 
algae,  clouds  of  pollen  and  spores  from  the  primeval  forests 
have  been  deposited  :  these  "  rains  of  sulphur  "  also  sank  in 
the  peaty  water  and  became  saturated  with  bitumen,  hence 
the  so-called  spore  and  pollen  coals. 

We  ought  not  to  be  too  ready  to  reject  the  bacterial  theories 
of  the  formation  of  the  various  coals,  especially  should  it  turn 
out  to  be  impossible  to  explain  without  microbes  the  formation 
of  the  bitumen  which  impregnates  them.  The  small  lower 
algae  are  rich  in  fats  and  capable  of  yielding  petroleum  bodies 


GENERAL   FUNCTIONS   OF   MICROBES        9 

by  distillation.  The  problem  of  the  origin  of  coal  has  therefore 
some  relation  to  the  problem  of  the  origin  of  petroleum.  The 
decomposition  of  fatty  substances  commences  with  their 
saponification,  which  splits  them  into  fatty  acids  and  glycerine, 
and  glycerine  is  a  good  food  medium  for  various  bacteria. 

The  moulds  decompose  fatty  acids  into  carbonic  acid  and 
water.  Fats  resist  decomposition  longer  than  other  carbo- 
hydrates, and  longer  than  nitrogenous  substances ;  it  is 
from  this  cause  that  the  proportion  of  fat  increases  in  a  cheese 
which  has  been  kept,  or  in  a  dead  body  which  is  decomposing. 
The  '  adipocere '  is  the  final  stage  in  an  animal  body  left  entirely 
to  nature- 
Ill.  Putrefaction  of  albuminous  materials. 

Putrefaction  returns  to  the  soil  the  nitrogen  which 
makes  up  1 5  per  cent,  of  the  proteins  of  animal  tissues. 

Since  plants  draw  their  nitrogen  from  the  soil  in  the  form  of 
nitrates  and  salts  of  ammonia,  it  is  necessary  for  the  complex 
protein  molecule  to  be  broken  down  so  as  to  supply  finally 


FIG.  2. — Ammoniacal  fermentation  Fig.  3.  —  Urobacillus  of 

of  urine  :   Urocococcus  of  Pasteur.  Duclaux. 

nitrogen  in  the  form  of  ammonia  and  nitric  acid.  Later 
green  plants  and  finally  animals,  raise  this  mineral  nitrogen  to 
the  level  of  organic  nitrogen. 

Already  during  life  animals  discharge  nitrogen  with  their 
excretions  in  the  form  of  urea,  uric  acid  and  hippuric  acid. 
The  Urobacteria  (there  are  about  a  hundred  species  known) 


10  MICROBES   AND  TOXINS 

transform  the  urea,  by  means  of  a  urease  which  they  secrete, 
into  carbonate  of  ammonia.  Hippuric  acid  is  transformed 
into  benzoic  acid  and  glycocoll,  and  finally  into  ammonia. 

The  putrefactions  of  albuminous  materials  in  nature  are 
never  simple,  that  is  to  say  carbohydrates  almost  always 
accompany  the  proteins  ;  even  meat  contains  a  little  sugar. 
Putrefactions  therefore  are  almost  always  mixed  fermentations. 

Pasteur  thought  that  all  putrefaction  was  the  work  of 
anaerobes.  He  discovered  the  vibrion  septique,  an  anaerobic 
bacillus  capable  of  decomposing  proteins.  Later,  the 
anaerobes  were  neglected  and  it  was  thought  that  various 
aerobes,  in  particular  Proteus^  were  the  principal  agents  of 
putrefaction.  But  after  the  study  of  fetid  suppurations  had 
drawn  attention  to  the  presence  of  anaerobes  (Veillon),  the 
idea  rose  again  that  they  too  must  play  a  part  in  putrefaction, 
and  the  methodical  study  of  this  subject  was  recommenced, 
chemical  analysis  going  hand  in  hand  with  bacteriological 
examination. 

In  their  experiments,  Tissier  and  Martelly  followed 
for  months  the  events  which  took  place  in  flasks  into  which 
meat  had  been  put  and  allowed  to  putrefy  either  with  or 
without  access  of  air.  Meat  taken  from  the  slaughter-house  as 
fresh  as  possible  contains  all  the  germs  necessary  and 
sufficient  for  putrefaction,  and,  as  it  contains  carbohydrates,  it 
is  a  mixed  putrefaction  which  occurs. 

There  are  two  phases.  In  the  first  the  sugar  and  the 
proteins  are  attacked  by  mixed  ferments,  that  is  to  say,  by 
microbes  which  decompose  at  the  same  time  both  proteins  and 
sugar,  proteolytic  and  saccharolytic — (peptolytic  is  the  term 
applied  to  the  ferments  which  attack  protein  only  after 
its  reduction  to  peptone).  In  the  second  phase  the  protein 
and  its  products  are  attacked  by  ferments  which  are 
proteolytic  or  peptolytic  pure  and  simple,  not  saccharolytic. 

But  between  these  two  phases  a  critical  turning-point  occurs: 
the  decomposition  of  the  sugars  produces  an  acidity  sufficient 
to  stop  putrefaction.  The  "antagonistic  force"  which 
Bienstock  observed  in  his  investigations  on  a  putrefactive 


GENERAL  FUNCTIONS   OF  MICROBES      11 

bacterium  the  B.putrificus,  is  nothing  but  this  acidity.  A  piece 
of  meat,  as  is  well  known,  keeps  excellently  in  unboiled  milk, 
because  the  milk  on  fermenting  produces  lactic  acid,  which 
protects  the  proteins  of  the  meat  from  putrefaction.  That  is 
why  the  housewife  when  she  salts  down  meat,  does  not  forget 
to  add  some  vinegar,  i.e.,  acid.  It  would  seem  then  that 
putrefaction  ought  to  cease  after  the  action  of  the  mixed 
ferments. 

If  it  does  not  stop,  it  is  because  there  is  not  sufficient  sugar. 
The  limit  of  acidity  for  the  pure  proteolytic  ferments  has  not 
been  reached.  Further,  owing  to  the  decomposition  of  the 
albumins  already  begun,  there  appears  a  base,  ammonia,  which 
neutralises  the  acidity,  and  the  pure  proteolytic  ferments  can 
begin  their  action.  It  is  the  anaerobes  which  break  down  the 
protein  molecule  and  produce  putrefaction;  but  they  cannot 
do  without  the  auxiliary  aerobes ;  thanks  to  the  aerobes  which 
hasten  the  production  of  ammonia  and  the  alkalinisation  of  the 
medium,  putrefaction  passes  successfully  through  the  crisis  of 
the  antagonistic  acidity. 

In  the  putrefaction  of  milk,  two  analogous  phases  succeed 
each  other.  Milk  being  richer  in  sugar  than  meat,  the  acidity 
developed  in  the  first  phase  is  greater  and  the  crisis  more 
difficult  to  surmount.  To  surmount  it,  more  powerful  fer- 
ments are  required  than  the  aerobes  which  produce  the 
ammonia  in  the  putrefaction  of  meat :  these  are  the  fungi 
(Oidium  lactis^  Rhizopus nigricans )  and  the  yeasts,  which  destroy 
the  acids,  consume  the  milk-sugar,  and  attack  the  casein. 
The  fungi  prepare  the  way  for  the  pure  ferments,  which  then 
carry  out  the  second  phase  of  putrefaction. 

When  an  animal  dies,  all  the  microbes  necessary  for  its 
putrefaction  are  present  in  its  intestine.  They  invade  the 
tissues  and  carry  out  on  a  larger  scale  what  we  see  on  a  small 
scale  in  the  experimental  flasks.  The  worm  of  the  grave  is  an 
old  poetical  image  long  out  of  date.  The  real  destroyer  is  the 
microbe. 

It  is  a  fact  well  established  to-day,  that  in  a  normal  intestine, 
the  bacteria  of  putrefaction  are  capable  of  vegetative  existence, 


12  MICROBES   AND  TOXINS 

and  that  our  digestion  is  accompanied  by  a  commencement  of 
putrefaction, — and  of  intoxication.  The  aerobic  bacteria  of 
the  Proteus  group  can  produce  putrefaction  there,  but  it  is 
chiefly  the  anaerobes,  as  was  Pasteur's  opinion,  which  do  this. 
There  have  been  found  in  the  human  intestine  all  the  most 
important  of  these,  the  B.  putrificus,  the  B.  sporogenes,  and 
the  B.  perfringens.  The  anaerobic  flora  of  the  intestine  does 
not  differ  much  from  the  anaerobic  flora  of  the  putrefying  meat 
in  Tissier's  experiments.  These  microbes  produce  poisons 
which  are  the  true  source  of  auto-intoxications  (Metchnikoff). 
For  life  in  general,  putrefaction  is  necesssary  to  permit  of  the 
circulation  of  nitrogen  in  nature  :  but  our  own  particular 
interest  demands  that  putrefaction  should  not  begin  too  soon, 
i.e.,  in  our  intestine,  and  mask  itself  under  an  appearance  of 
perfect  health. 

To  combat  the  intestinal  putrefaction,  it  is  necessary  to 
adopt  a  diet  capable  of  producing  in  us  that  limit  of  acidity 
which  induces  the  crisis  separating  the  two  phases  of  putre- 
faction of  meat  or  milk,  and  of  arresting  decomposition  in  our 
bodies  at  the  end  of  the  first  phase.  Therefore,  we  ought  to 
eat  carbohydrates  and  sugars,  and  so  alter  the  conditions 
in  our  intestine  as  to  favour  the  lactic  ferments. 

IV. — Nitrification  and  Denitrification. 

Nitrates  represent  the  form  of  nitrogen  preferred  by  the 
plants,  and  it  has  long  been  known  that  the  ammonia  set  free 
in  putrefaction  becomes  oxidised  in  the  soil,  the  ammonium 
salts  being  transformed  into  nitrates.  This  is  the  process 
of  nitrification  and  is  carried  on  by  bacteria. 

It  was  long  thought  that  the  ammoniacal  salts  were  oxidised 
in  contact  with  the  soil  by  the  direct  action  of  atmospheric 
oxygen,  in  the  same  way  that  certain  chemical  combinations 
can  take  place  on  contact  with  porous  substances.  But  chalk 
and  sand,  which  ought  to  act  as  porous  substances,  cannot 
take  the  place  of  earth.  Pasteur  perceived  with  his  peculiar 
intuition  that  the  lower  plant  must  play  a  part  in  nitrification. 


GENERAL   FUNCTIONS   OF   MICROBES      13 

That  nitrification  is  the  work  of  living  creatures  has  been 
proved  by  the  celebrated  experiment  of  Schlcesing  and  Miintz. 
If  a  cylinder  is  filled  with  cultivated  soil,  ammonia  poured  on 
the  top  appears  at  the  bottom  as  nitrate  of  lime.  But  this 
transformation  no  longer  occurs  if  the  earth  is  previously  heated 
to  100°  C,  or  if  it  is  impregnated  with  the  vapour  of  chloroform 
or  of  carbon  bisulphide,  i.e.t  if  the  living  organisms  that  it 
contains  are  killed  or  paralysed.  The  soil  recovers  its  activity 
when  the  paralysing  vapours  have  been  removed  by  passing 
through  a  current  of  air. 

Nitrification  is  a  process  of  two  stages,  and  is  carried  out  by 
two  species  of  bacteria,  each  with  its  own  function.  In  the 
first  phase,  the  ammoniacal  salts  are  transformed  into  nitrites 
by  the  microbes  known  as  the  nitrous  ferment,  e.g.  Nitroso- 
monas  of  Europe,  Nitrosomonas  of  Java,  and  the  Nitrosococcus 
of  America  (Mexico  and  Brazil).  In  the  second  phase  the 
nitrites  are  turned  into  nitrates  by  the  nitro-bacterium  or  nitric 
ferment,  the  Nitrobacter  of  Winogradsky  (figs.  4-6). 

Neither  the  nitrous  nor  the  nitric  ferments  develop  in 
presence  of  organic  matter.  The  activity  of  the  nitrous 
ferment  is  arrested  by  0*3  per  cent,  of  glucose,  peptone  or 
asparagin.  The  nitric  ferment,  less  sensitive,  is  stopped  by 
0-3  per  cent,  of  glucose,  1^25  per  cent,  of  peptone  or  i  per 
cent,  of  asparagin.  The  former  is  inhibited  by  1*5  per  cent., 
the  latter  by  3  per  cent,  of  sodium  acetate. 

Now  we  are  accustomed  to  the  idea  that  bacteria  live  on 
organic  matter.  Yet  the  bacteria  of  nitrification  must,  as  we 
have  seen,  get  their  nourishment  elsewhere.  They  differ  from 
other  bacteria  in  being  capable  of  taking  up  carbon  by  decom- 
posing carbonates,  in  being  aerobic,  and  in  behaving  like  chloro- 
phyllous  plants.  They  are  not  saprophytes  in  the  same  sense 
as  the  majority  of  bacteria. 

In  the  laboratory,  the  two  phases  of  nitrification  can  be 
demonstrated  separately  by  pure  culture.  But  in  nature  they 
are  simultaneous.  Under  laboratory  conditions  ammonia 
exerts  an  inhibiting  action  on  the  nitric  ferment,  whereas  in 
nature  both  the  actions  can  occur  in  presence  of  quantities  of 


14  MICROBES   AND  TOXINS 

ammonia  frequently  very  considerable.  Schloesing  has  shown 
that  in  the  soil  ammonia  does  not  prevent  the  nitric  ferment 
from  acting.  These  apparent  contradictions  between  theory 
and  practice  are  capable  of  explanation.  Ammonia  prevents 
the  development  of  the  nitric  ferment  but  scarcely  acts  at  all  on 
the  same  ferment  in  the  adult  state.  The  soil  in  nature  being 
populated  by  the  adult  ferment,  the  inhibiting  action  of 
ammonia  is  much  more  limited  than  in  an  experimental 
flask  (Boullanger  and  Massol). 

The  nitrous  ferment  will  stand  neither  an  excess  of  ammonia 
nor  an  excess  of  its  own  product,  the  nitrite  of  magnesium, 
The  nitric  ferment  ceases  to  act  when  too  much  nitrite  has 


VK3U%<>     VX>    ^ 

**%/£ 


FIG.  4. — Nitrous  ferment  FIG.  5. — Nitric  ferment :  the  nitro-bacte- 

from  Gennevilliers.  rium  from  Quito  (after  Kayser). 

been  produced  around  it  and  when  it  has  itself  developed  a 
certain  quantity  of  nitrates.  Thus  each  demands  suitable 
proportions  both  of  the  primordial  material  and  of  the  products. 
It  is,  no  doubt,  to  maintain  these  favourable  conditions  that 
we  have  the  denitrifying  microbes  which  partially  undo  the 
labour  of  the  nitrifying  organisms.  They  return  the  nitrates 
and  nitrites  to  the  condition  of  ammonia,  liberating  protoxide 
of  nitrogen,  dioxide  of  nitrogen,  or  simply  nitrogen.  Wheat 
straw,  the  straw  of  maize  and  of  lucerne  and  oilcakes  contain 
denitrifying  bacteria.  Animal  excrement  also  contains  them, 
for  soil  to  which  cow  dung  is  added  loses  part  of  its  nitrates. 
When  soil  is  treated  with  nitrate  of  soda  too  soon  after 
receiving  farmyard  manure  it  loses  nitrogen.  The  denitrifying 
organisms  act  best  in  presence  of  the  excess  of  organic  matter 


GENERAL   FUNCTIONS   OF  MICROBES      15 


which  is  so  prejudicial  to  nitrification.  Probably  the  two 
functions  balance  and  regulate  each  other.  The  nitrogen 
liberated  by  the  denitrifying  bacteria  is,  however,  not  lost ;  it 
may  be  taken  up  by  the  bacteria  which  fix  nitrogen.  It  is 
nevertheless  true  that  farmers  ought  to  be  on  their  guard 
against  the  denitrifying  organisms  and  avoid  putting  on  the 
soil  farmyard  manure,  especially  if  fresh,  along  with  nitrates. 
Formerly  in  the  manufacture  of  saltpetre  cultivated  soil  was 


$ 
»« 


FIG.  6. — Nitrifying  bacteria  (after  Winogradsky). 

A.  Nitrous  ferment  from  Zurich. 

B.  The  same  in  the  motile  form  with  flagella. 

C.  Nitrous  ferment  from  Kazan. 

D.  Nitrous  ferment  from  Java,  motile  cells  and  groups  of  cells. 

freely  sprinkled  with  urine.  If  the  deposits  of  nitrate  in  Chili 
and  Peru  should  some  day  be  exhausted,  it  would  not  be 
impossible  to  make  it  with  the  help  of  bacteria  (not  to  speak  of 
the  electrical  methods  which  have  already  been  employed). 
Saltpetre  is  produced  everywhere  when  suitable  conditions  of 
moisture  and  organic  matter  are  given.  The  saltpetre  of 


16  MICROBES  AND  TOXINS 

cellars  is  nitrate  of  lime  which  has  risen  from  the  soil  through 
the  walls  by  capillarity  and  has  undergone  evaporation. 

Long  ago  Miintz  prepared  artificial  nitre  beds  which  furnished 
eight  grams  of  nitrate  daily;  in  recent  experiments  he  and 
Laine  have  obtained  such  a  yield  that  they  find  themselves 
capable  of  preparing  by  means  of  nitrifying  organisms  all  the 
saltpetre  for  the  powder  necessary  to  defend  the  nation  in  war. 
Distributed  over  beds  of  peat  of  two  metres  thickness  on  the 
top  of  a  layer  of  clinker,  the  nitrifying  bacteria  are  capable  of 
producing,  for  each  25  acres  of  area,  1,500  tons  of  nitrate  per 
day,  after  a  starting  period  of  one  month  at  most  to  get  the 
beds  into  working  order;  in  five  days,  that  is,  7,500  tons,  or 
the  annual  requirement  of  powder  for  the  army.  It  is  easy  to 
calculate  what  750,000  acres  of  peat  bogs  in  France  could 
produce  if  necessary. 

The  purification  of  sewage  is  one  of  the  greatest  tasks  which 
burden  the  hygienists  of  large  towns.  Broad  irrigation  followed 
by  cultivation  demands  much  land  and  is  not  quite  safe  except 
when  employed  solely  for  the  growth  of  forage,  not  for  market- 
gardening.  Hence  it  is  gradually  giving  place  to  the  biological 
method  of  purification,  an  intensive  process  carried  on  in 
small  space,  and  here  again  by  bacteria. 

We  shall  not  enter  here  upon  the  details  of  its  application. 
In  principle,  complete  biological  purification  goes  on  in  two 
phases :  a  phase  of  anaerobic  fermentation  in  the  septic  tank 
and  a  phase  of  aerobic  fermentation  in  the  bacterial  beds. 

In  the  septic  tank,  into  which  the  sewage  must  be  run 
gently — so  as  not  to  carry  in  the  air  which  would  favour 
aerobic  fermentation — in  which  it  must  be  allowed  to  circulate 
quietly,  and  to  stay  at  least  twenty-four  hours,  the  disintegration 
takes  place  both  of  hydrocarbons  and  proteins  under  the 
agency  of  legions  of  bacteria  which  secrete  all  sorts  of  diastases. 
The  sludge  dissolves  and  the  resultant  product  can  be  sub- 
mitted to  the  action  of  the  nitrifying  agents. 

The  experimental  control  of  the  ferment  activity  in  the 
septic  tank  can  be  carried  out  by  comparing  the  action  on 
coagulated  egg-white,  meat,  raw  or  cooked,  fats,  paper,  &c. 


GENERAL  FUNCTIONS   OF  MICROBES      17 

In  six  weeks  100  grams  of  egg-white  leave  only  one  gram  of 
residue,  whereas  in  stagnant  sewage  there  remain  76  grams, 
and  in  running  water  83  grams.  In  three  weeks  meat  loses 
almost  50  per  cent,  of  its  weight,  and  in  six  weeks  96  per  cent. 
The  body  of  an  animal,  immersed  in  it  is  for  long  protected 
by  the  layer  of  subcutaneous  fat ;  but  cleaned  cartilage  and 
tendon  lose  in  five  weeks  65  to  99  per  cent,  of  their  weight ; 
even  wool  and  feathers  decompose. 

As  regards  hydrocarbons,  the  fats  are  slowly  split  into  fatty 
acids  and  glycerine.  Cabbages  and  potatoes  are  almost 
completely  destroyed  in  six  weeks.  A  hempen  rope  which, 
after  five  weeks  of  immersion  in  stagnant  sewage  or  running 
water,  could  still  bear  the  weight  of  12  kilograms,  broke  under 
15  grams  after  the  same  period  of  immersion  in  the  septic 
tank.  After  three  weeks  newspaper  begins  to  dissolve  libera- 
ting bubbles  of  gas.  It  is  quite  wrong  to  consider  the  septic 
tank  as  operating  like  a  simple  settlement  tank  :  it  is  rather  a 
sort  of  crucible  in  which  the  powerful  microbes  melt  and 
disintegrate  the  most  resistant  organic  matter.  The  septic 
tank  liberates  various  gases,  methane,  hydrogen,  nitrogen  and 
carbonic  acid,  a  cubic  metre  of  sewage  furnishing  from  40  to 
70  litres  of  gas. 

The  aerobic  phase  of  the  purification  is  accomplished  by 
bacterial  beds,  into  which  the  organic  matter,  already  dissolved 
and  transformed  into  ammoniacal  compounds  is  discharged. 
What  takes  place  in  them  is  an  intense  nitrification  carried 
on  by  the  same  microbes  which  in  arable  soil  transform  the 
ammonia  into  mineral  salts,  nitrites  and  nitrates. 

The  bacterial  bed  consists  of  a  thick  layer  of  clinker  or  slag, 
and  is  filled  with  sewage  for  periods  of  an  hour  and  a  half  to 
two  hours  separated  by  intervals  of  four  to  six  hours.  Contact 
may  be  renewed  if  necessary  two  or  three  times  by  passing  the 
effluent  through  a  second  or  third  bacterial  bed.  During 
contact  the  organic  matter  fixes  itself  on  the  clinker,  while 
during  the  aeration  period  it  is  oxidised  by  the  ferments  which 
take  up  the  necessary  oxygen  from  the  air. 

The  nitrification  is  balanced  in  the  bacterial  beds  by  a 

c 


18  MICROBES   AND  TOXINS 

denitrification  as  in  cultivated  soil.  Nitrifying  and  denitrifying 
ferments  can  live  side  by  side;  the  latter  only  interfere  with 
the  object  desired  when  there  is  present  an  excess  of  hydro- 
carbons. 

A  town  of  500,000  inhabitants,  furnishing  6  million  gallons 
of  waste  water,  could  replace  the  682  acres  required  for  broad 
irrigation,  or  the  15  acres  required  for  the  bacterial  beds  with 
slags  and  clinker,  by  2*5  acres  of  bacterial  bed  built  on  peat 
(Calmette). 

The  biological  purification  returns  the  nitrogen  to  nature  in 
the  form  of  nitrate,  although  a  certain  quantity  is  lost  in  the 
form  of  gas.  There  are  other  bacteria,  however,  capable  of 
taking  up  nitrogen  from  the  air  and  re-introducing  it  into  the 
cycle  of  animal  and  vegetable  life. 

V. — Fixation  of  Atmospheric  Nitrogen  in  the  Soil. 

The  life  of  both  animal  and  vegetable  species  depends  on  the 
stock  of  nitrogen  retained  by  the  soil.  Although  the  earth 
acquires  nitrogen  from  putrefying  processes  it  loses  nitrogen 
also,  discharged  in  the  gaseous  condition;  some  is  lost  also 
during  denitrification,  and  in  percolating  water  which  robs 
Uncultivated  soil  of  as  much  as  40  kilos,  of  nitrogen  per  acre 
per  annum.  Floods  also  carry  off  the  nitrates,  and  after  the 
floods  of  1896,  Schloesing  calculated  that  the  Seine  carried  off 
about  5  milligrams  of  nitric  acid  per  litre,  at  the  rate  of 
800,000  litres  per  second ;  the  total  nitric  acid  lost  amounted 
to  350,000  kilos,  per  twenty-four  hours,  equal  to  650,000  kilos, 
of  saltpetre.  The  rivers  pour  this  nitrogen  into  the  sea. 

And  yet  in  spite  of  these  losses  the  soil  retains  its  nitrogen. 
Nay  more,  it  accumulates  it.  The  soil  of  forests  is  never 
manured  and  the  woodcutters  carry  off  a  great  quantity  of 
nitrogen  with  the  wood ;  yet  the  soil  there  remains  fertile.  In 
the  hill  pastures,  flocks  are  feeding  all  the  summer  and 
furnishing  us  with  nitrogen  in  the  form  of  milk,  cheese  and 
meat :  and  yet  the  soil  of  these  natural  fields  contains  quantities 
of  nitrogen  greater  than  is  found  in  soil  ploughed  and  copiously 


GENERAL  FUNCTIONS   OF   MICROBES      19 

manured,  /.*.,  from  5  to  9  milligrams  of  combined  nitrogen 
per  kilo,  of  soil.  Finally,  the  crops  remove  from  the  soil 
much  more  nitrogen  than  the  manuring  supplies  :  the  difference 
varying  with  the  rotation  of  the  crops  from  1*5  to  400  kilos, 
per  acre.  Where  does  this  nitrogen  come  from  ? 

It  can  only  come  from  the  inexhaustible  reservoir  of  the 
atmosphere. 

Rain  water  carries  into  the  earth  the  ammonia  which  has 
evaporated  from  it  and  the  oxidised  compounds  of  nitrogen 
which  form  during  thunderstorms.  But  these  gains — about 
1*5  kilos,  per  acre  per  annum — are  quite  insufficient  to  com- 
pensate for  the  losses  occasioned  by  drainage  and  cultivation. 
Plants  must  take  up  not  only  the  ammonia  and  the  nitrates 
of  the  atmosphere  supplied  by  rain,  but  also  uncombined 
nitrogen,  the  free  nitrogen  of  the  air. 

Cultivated  soil,  kept  moist  and  exposed  to  the  air,  fixes 
atmospheric  nitrogen  (Berthelot).  This  fixation  does  not  occur 
if  the  earth  has  been  sterilised  by  heating  to  120°  C.  Living 
creatures  must  therefore  be  at  work  in  this. 

These  workers  are  the  bacteria  of  the  soil  which  are  found 
to  a  depth  of  one  foot.  They  are  also  found  in  the  sea, 
especially  in  the  neighbourhood  of  algae.  They  are  more 
abundant  in  soil,  the  better  it  is  aerated  and  cultivated.  They 
include  anaerobes  (Clostridium  pasteurianum)  and  aerobes 
(genus  Azotobacter :  Pseudomonas  leuconitrophilus).  Their 
organic  food  in  soil  as  in  laboratory  cultures  is  carbohydrate, 
glucose,  saccharose,  levulose,  dextrine,  mannite  and  other 
sugars;  butyrates,  lactates  and  acetates;  their  mineral  food 
consists  of  salts  of  lime  and  phosphates.  The  Azoto- 
bacter chroococcum  will  only  develop  in  soil  containing  at  least 
o'i  per  cent,  of  lime. 

It  is  not  exactly  known  how  bacteria  fix  nitrogen.  They 
build  it  up  into  their  substance  and  liberate  it  when  they  are 
destroyed.  Doubtless  also  they  build  up  nitrogenous  com- 
pounds which  are  taken  up  by  the  nitrifying  bacteria. 

Nitrogen  can  only  be  thus  combined  on  condition  that  the 
bacteria  are  supplied  with  energy  in  the  form  of  carbohydrate 

C    2 


20  MICROBES   AND  TOXINS 

food.  No  hydrocarbons,  no  bacterial  activity,  no  fixation  of 
nitrogen.  To  fix  a  gram  of  nitrogen,  experiments  show,  100 
to  200  grams  of  glucose  are  required. 

Just  as  we  do  not  know  all  about  a  disease  when  we  know 
its  microbe,  so  it  is  not  sufficient  to  have  simply  the  bacteria 
which  fix  free  nitrogen  in  order  to  enrich  at  will  a  soil  with 
nitrogen.  Nature  is  not  the  laboratory.  The  inoculation  of 
poor  soil  with  these  beneficent  bacteria  produces  scarcely  any 
increase  in  the  crops.  To  excite  bacterial  multiplication  it 
would  be  necessary  to  distribute  sugars  over  the  soil — 14 
kilos.,  it  is  calculated,  to  fix  the  nitrogen  to  which  corresponds 
i  kilo,  of  nitrate.  Nitrogen  at  such  a  price  is  far  from  cheap. 
Besides,  the  hydrocarbons  favour  the  denitrifying  action  of  the 
microbes  which  results  in  a  loss  of  nitrogen.  The  useful 
germs  are  already  present  in  good  soil ;  it  is  literally  the  soil 
itself  which  must  be  altered,  by  adding  marl,  by  tilling,  drainage, 
and  all  the  operations  which  change  its  physical  properties. 

There  exist  certain  moulds  which  fix  the  nitrogen  of  the  air, 
for  example  the  Penidllia  and  the  Sterigmatocystes.  The  algae 
of  the  nostoc  group,  Chlorella^  Stichococcus  and  Cystococcus,  fix 
nitrogen  only  when  in  symbiosis  with  the  fixing  bacteria  :  it  is 
then  the  bacteria  which  fix  nitrogen  ;  the  algae,  as  green  plants, 
nourish  their  associated  bacteria  by  means  of  the  hydrocarbons 
which  they  synthesize  in  virtue  of  their  chlorophyll  activity. 
According  to  Beijerinck's  experiments  the  Cyanophyceae  are 
capable  of  taking  nitrogen  into  their  tissues  independently  just 
as  they  do  carbon.  But  if  all  the  algae  possessed  this  double 
function  they  would  be  so  powerfully  adapted  for  life  that  they 
must  have  long  ago  invaded  the  whole  universe. 

There  is  a  third  group  of  microbes  which  fix  nitrogen ;  but 
in  this  case  they  do  not  live  free  in  the  soil,  but  are 
confined  to  the  roots  of  Leguminosae. 

The  soils  which  are  found  to  be  most  rich  in  nitrogen  are 
those  in  which  there  have  been  longest  grown  crops  of  this 
family.  Georges  Ville  in  1852  thought  that  the  leguminosae 
took  up  oxygen  from  the  air.  He  sowed  leguminosae  in  sand 
which  had  been  washed  and  calcined,  thus  being  sterile  and 


GENERAL   FUNCTIONS   OF   MICROBES      21 

freed  from  nitrogen  compounds  ;  he  kept  them  in  an  atmosphere 
similarly  freed  from  all  compounds  of  nitrogen  ;  when  the  young 
plants  had  overcome  these  rather  unfavourable  conditions  they 
contained  at  the  end  of  the  experiment  more  nitrogen  than 
was  present  in  the  seeds. 

Malpighi  observed  long  ago — in  1687 — little  nodules 
on  the  roots  of  Leguminosae  whose  function  has  only  in 
modern  times  been  discovered  thanks  to  Pasteur.  These  little 
nodules  are  in  fact  crammed  with  bacteria  the  function  of  which 
is  to  fix  the  atmospheric  nitrogen.  The  plant  suffers  from  a 
malady  which  is  actually  beneficent  (Hellriegel  and  Wilfarth). 

Grown  in  a  sterile  soil  and  protected  from  the  germs  of  the 


FIG.  7. — Nodules  on  the  roots  of  Leguminosae  ( Viciafaba}. 

air  and  of  the  soil,  the  roots  of  Leguminosae  never  present 
these  nodosities.  If  this  soil  is  watered  with  a  suspension  of  the 
nodules  or  of  earth  in  which  Leguminosae  have  grown,  or  is 
simply  sprinkled  with  garden  soil,  the  Leguminosae  grow  in 
it  possessing  nodules,  but  if  these  suspensions  are  boiled  for  a 
sufficient  time  they  lose  this  property. 

It  is  possible  to  inoculate  the  plant  with  the  infection  by 
simply  pricking  with  a  needle  first  a  nodule  on  the  root  of  a 
normal  pea,  then  the  root  of  a  pea  which  has  been  grown 
aseptically  without  nodules ;  under  such  conditions  the  roots 
which  were  free  from  nodules  develop  them. 

The  bacteria  of  the  nodules  have  an  irregular  and  peculiar 
shape,  and  have  been  given  the  name  of  bacteroidia.  The 
best  known  is  the  B.  radicicola  of  Beijerinck.  Two  groups 


22  MICROBES   AND   TOXINS 

have  been  made  :  those  of  the  vetches,  clovers,  and  peas,  and 
those  of  the  beans  and  lupines.  Maze  distinguishes  those 
adapted  to  calcareous  soil,  the  '  calcicolae  '  from  those  adapted  to 
acid  soils,  the  '  calcifugae.'  They  are  aerobic,  and  pure  cultures 
can  be  obtained.  They  require  carbohydrate  food,  and  are 
not  fond  of  nitrates.  Grown  in  pots,  the  nodules  are 
encouraged  by  the  addition  of  chromium,  manganese,  nickel, 
and  cobalt.  About  100  grams  of  sugar  are  consumed  for  each 
gram  of  nitrogen  fixed.  The  bacteroidia  only  act  well  when 
their  food  supply  is  regulated  in  quantity  as  well  as  in  quality. 
The  broth  for  the  cultures  should  contain  i  per  10,000  at 


8. — Bacteria  and  Bacterioidia  from  the  nodules 
of  Leguminosse  (after  Beijerinck). 

least  and  i  per  3,000  at  most  of  combined  nitrogen  ;  the 
proportions  of  saccharose  should  lie  between  2  and  6  per 
cent. 

The  virulence  of  the  bacteroidia  varies  \  those  which  have 
undergone  several  passages  in  the  roots  attack  the  new  roots 
more  easily.  It  seems  that  the  weaker  bacteria  induce  a 
certain  immunity  in  the  rootlets  towards  the  more  virulent. 
A  root  already  bearing  nodules  does  not  produce  new  ones 
except  when  inoculated  with  very  virulent  bacteroidia.  On  the 
same  plant  the  recent  nodules  of  the  lateral  roots  contain 
bacteria  much  more  virulent  than  those  of  the  main  root. 


GENERAL  FUNCTIONS   OF   MICROBES      23 

The  bacteroi'dia  scattered  through  the  soil  are  attracted 
towards  the  young  roots  by  the  vegetable  carbohydrates  :  there 
is  a  positive  chemiotaxis. 

The  bacteroi'dia  and  the  plant  are  mutually  beneficial. 
The  plant  supplies  carbohydrate  food  to  the  bacteroi'dia,  which 
n  their  turn  supply  the  plant  with  nitrogen.  In  culture  a 
viscous  glairy  jelly  appears,  which  is  present  also  in  the 
nodules,  but  only  in  their  early  stage.  The  sap  rapidly  carries 
it  into  the  body  of  the  plant,  and  it  is  this  jelly  probably  which 
contains  the  nitrogenous  food  product.  Some  diastase 
probably  of  the  root  liberates  the  nitrogen  compound  from  the 
cells  of  the  bacteroi'dia. 

Nitrogenous  manures  are  expensive,  and  hence  the  idea 
has  arisen  to  improve  leguminous  crops  by  planting  them  in 
prepared  soil,  or  by  adding  soil  which  has  already  grown 
leguminous  plants.  Afterwards  the  attempt  was  made  to 
inoculate  the  soil  with  artificial  cultures  of  the  nodule 
bacteria.  The  process  is  analogous  to  the  injection  into  an 
animal  of  a  food  substance  or  a  drug:  it  is  agricultural 
bacteriotherapy . 

In  observing  nature,  not  from  the  point  of  view  of  any 
single  living  species,  man  or  animal,  but  throughout  the  whole 
of  her  operations,  one  sees  that  the  pathogenic  bacteria  are 
less  prominent  than  the  useful  ones — one  may  say  even  that 
all  bacteria  are  useful  and  are  only  injurious  by  accident. 
They  all  have  their  role  in  the  cycle  of  matter  and  only  destroy 
one  existence  to  prepare  for  another. 

To  perceive  the  function  of  the  useful  bacteria,  it  has  been 
necessary  to  study  all  the  fermentations  in  relation  to  both 
industry  and  food  supply :  wine,  beer,  cider,  vinegar,  cheese, 
bread,  sauer-kraut,  tobacco,  and  the  leather  of  our  shoes  are 
all  more  or  less  the  result  of  bacterial  operations.  Life  as  a 
whole  could  not  continue  without  bacteria ;  they  do  not  create 
life,  but  they  supply  it  with  the  necessary  material. 

We  may  ask  ourselves  now  if,  with  nature  thus  provided 
with  nutritive  material,  the  life  of  a  particular  organism  might 
be  possible  although  it  did  not  contain  microbes  in  itself. 


24  MICROBES   AND   TOXINS 

Logically  it  is  possible.  An  animal  without  microbes,  an 
aseptic  creature,  might  take  advantage  of  the  general  activity 
of  microbes  without  itself  being  open  to  these  peculiar  fermen- 
tations which  we  call  diseases. 

This  then  would  be  at  once  the  first  principle  of  hygiene, 
and,  as  it  were,  its  paradox. 


CHAPTER  II 

MICROBES  IN  THE  HUMAN  BODY  — LIFE  WITHOUT  MICROBES — 
THE  INTESTINAL  FLORA 

Microbes  in  and  on  the  bodies  of  animals  and  man  :  skin,  mucous  mem- 
branes, mouth,  stomach,  intestine. 

Life  without  microbes —Pasteur's  ideas — Aseptic  breeding — The  example 
of  the  bat,  Pteropus. 

The  intestinal  flora  of  man  :  quantity,  species,  and  variations  of  the 
microbes. 

Intestinal  putrefactions— The  flora  and  diet — Products  of  putrefaction  and 
auto-intoxication. 

Associations  and  antagonisms— Cholera — Experiments  of  MetchnikofT, 
Bienstock,  Tissier — Principles  of  intestinal  bacteriotherapy — Sour 
milk  ;  culture,  pure  and  mixed,  of  the  lactic  ferments. 

Is  the  large  intestine  useless? — Is  the  intestine  permeable  to  microbes? — 
Investigations  on  anthracosis  and  tuberculosis — The  defensive  powers 
of  the  mucous  membrane. 

The  Microbial  Flora  in  Man   and   Animals. — We 

have  seen  what  innumerable  legions  of  bacteria  are  at  work  in 
the  universe.  Let  us  now  proceed  from  the  contemplation  of 
the  macrocosm  to  the  microcosm,  as  represented  by  our  own 
bodies. 

There  is  scarcely  a  living  being  which  is  not  crowded  with 
microbes.  Living  as  we  do  in  a  universe  where  they  swarm, 
how  could  we  avoid  having  them  both  within  and  without  ? 

Normally,  we  enter  the  world  free  from  microbes.  But  from 
the  first  moment  after  birth  they  begin  to  settle  on  our  skin 
and  on  our  mucous  membranes.  Our  mother's  first  touch 
communicates  them  to  us,  even  before  she  has  given  us  the  first 
drops  of  her  milk.  They  penetrate  the  nose,  the  mouth,  and 

25 


26  MICROBES   AND   TOXINS 

the  lungs  with  our  first  respiratory  movements  and  our  first 
cries ;  they  are  deposited  on  our  skin  from  our  first  bath  and 
our  first  swaddling-clothes.  Even  four  hours  after  birth  -  and 
invariably  between  the  tenth  and  the  seventeenth  hour  of  life 
— they  have  already  reached  the  intestine. 

Our  skin  is  inhabited  chiefly  by  round  bacteria,  such  as 
streptococci,  and  especially  staphylococci ;  these  latter  are  most 
abundant,  and  there  are  several  species  of  them,  some 
commoner  than  others.  They  do  not  inhabit  so  much  the 
smooth  surface  of  the  epidermis  as  the  hair  follicles  and  their 
adnexa,  the  sebaceous  glands,  which  are  regular  dens  of 
bacteria.  Even  the  cleanest  persons,  however  little  oily  their 
skins,  have  only  to  press  between  the  fingers  a  little  fold  of  skin 
on  the  end  or  at  the  side  of  the  nose,  to  squeeze  out  what 
looks  like  a  little  worm,  but  which  is  nothing  else  than  a  colony 
of  staphylococci. 

As  a  matter  of  fact,  however  healthy  the  skin,  it  is  more  or 
less  inhabited  by  bacteria,  which,  however,  in  general,  remain 
harmless ;  the  skin  is  not  infected.  But  given  at  some  point  a 
lesion,  a  boil  or  pustule,  where  bacteria  have  multiplied,  the 
microbes  of  these  foci  spread  themselves  over  all  the  cutaneous 
surface,  even  to  the  most  distant  parts.  "  On  the  surface  of 
healthy  skin,  the  diffusion  of  germs  takes  place  around  the 
original  lesion  for  a  great  distance,  and  with  an  abundance  and 
continuity  such  as  is  only  paralleled  by  the  law  of  conservation 
of  widely  distributed  species.  An  analogous  case  is  that  of 
plants  covering  square  miles  of  country  round  with  their 
pollen  or  trjeir  winged  seeds"  (Sabouraud).  Hence,  the  best 
way  to  avoid  cutaneous  infections  is  to  preserve  intact  the 
epidermis,  and  not  to  weaken  by  the  abuse  of  antiseptics  its 
protecting  cells,  which  endeavour  to  maintain  the  integrity  of 
the  skin  against  bacteria. 

The  mucous  membranes,  warmer  and  moister  than  the  skin, 
form  a  better  soil  for  microbes.  There  is  in  the  conjunctiva 
of  the  eye,  among  others,  a  little  bacillus,  resembling  the 
bacillus  of  diphtheria,  which  is  found  from  the  first  hours  of  life 
(Morax),  In  the  cavities  of  the  nose  and  pharynx,  there  are 


MICROBES    IN   THE  HUMAN  BODY         27 

not  only  the  common  bacteria,  the  streptococci  and  staphylo- 
cocci,  but  also  pathogenic  bacteria,  which  take  shelter  there 
and  remain  latent  until  an  opportunity  offers  to  multiply ; 
examples  are  the  diphtheria  bacillus,  the  microbe  of  pneumonia, 
and  that  of  cerebro-spinal  meningitis. 

In  the  deeper  respiratory  passages,  there  are  few  microbes ; 
there  they  may  be  numbered  by  units.  But  it  is  proved  that 
the  tubercle  bacillus  can  penetrate  with  the  inspired  air  right 
to  the  bottom  of  the  pulmonary  alveoli. 

The  mouth,  the  vestibule  of  the  digestive  tube,  contains 
already  a  large  proportion  of  the  microbial  species  which 
populate  the  stomach  and  intestine,  e.g.,  staphylococci  and 
streptococci,  resembling  those  of  the  skin,  bacilli,  aerobic 
and  anaerobic,  resembling  those  found  both  in  the  healthy 
intestine  and  in  the  intestine  and  appendix  in  disease,  and  finally 
a  whole  flora  peculiar  to  the  mouth  which  plays  a  part  in  dental 
caries. 

The  stomach  being  acid  suits  moulds  and  yeasts  better  than 
bacteria.  However,  about  thirty  species  of  bacteria  have  been 
described  in  it  (Coyon),  several  of  which  have  attracted  special 
attention  because  of  the  idea  that  they  might  favour  or  inhibit 
the  penetration  of  certain  pathogenic  bacteria  into  the  intestine. 
'  All  the  cavities  and  recesses  of  the  human  body  deserve 
study  from  the  point  of  view  of  their  flora.  Many  species  have 
been  seen  but  are  not  yet  well  known,  because  we  do  not  yet 
know  how  to  cultivate  them  artificially. 

The  flora  hitherto  most  studied  is  that  of  the  intestine.  It 
is  also  the  most  important.  The  intestine  is  the  great  laboratory 
of  digestion,  and  at  the  same  time,  unfortunately,  of  putrefac- 
tions, the  products  of  which  are  absorbed  by  the  body. 
Hence  one  may  say  that  man,  like  other  animals,  is  dependent 
on  his  belly,  and  no  system  of  therapy  is  justified  in  neglect- 
ing it.  Just  as  the  soil  outside  of  us  is,  in  nature,  the  great 
microbial  reservoir,  so  in  us  the  great  reservoir  of  microbes  is 
our  intestine. 

Under  the  impulse  of  Metchnikoff,  the  intestine  has  with 
justice  become  the  great  field  of  study  and  experiment  in  those 


28  MICROBES  AND  TOXINS 

problems  of  nutrition  which  affect  most  of  all  the  general 
health,  the  individual  development,  and  the  evolution  of  man. 
That  is  why  we  pay  special  attention  here  to  the  intestinal 
microbes  as  being  the  most  important  of  all  those  which 
inhabit  the  bodies  of  man  and  animals. 

The  first  problem  in  this  question  is  the  possibility  of  life 
without  microbes. 

Life  without  Microbes. — Life  in  general,  as  we  know 
it  on  the  surface  of  this  earth  of  ours,  is  impossible  without 
the  action  of  the  micro-organisms.  As  some  philosophers 
have  well  said,  "life  is  but  a  little  mould  growing  on  the 
surface  of  a  rather  moist  planet."  But  is  it  possible  for  certain 
individuals  to  live  free  from  bacteria,  although  of  course 
depending  on  the  conditions  produced  by  bacteria,  and  paying 
tribute  to  bacteria  by  reason  of  the  fermentations  to  which 
they  owe  their  origin  and  nourishment  and  by  which  one  day 
they  are  doomed  to  be  dissolved  ? 

Plants  have  bacteria  both  around  their  roots  and  in  their 
tissues.  Animals  have  thousands  of  millions  in  their  digestive 
tubes.  The  microbial  world  not  only  surrounds  them  but 
is  in  them.  In  our  intestine  there  proceed  fermentations 
from  which  we  can  absorb  the  products.  Is  this  to  our 
advantage  or  is  it  not  ? 

It  seems  that  plants  derive  only  benefit.  The  bacteria  prepare 
for  them  their  food.  A  sterile  seed  made  to  sprout  in  a  sterile 
nutritive  fluid,  such  as  milk,  can  use  neither  casein  nor  sugar 
nor  starch ;  it  secretes  neither  rennin  nor  casease,  nor  sucrase, 
nor  amylase.  There  is  amylase,  it  is  true,  in  the  cotyledons  of 
the  germinating  seed,  but  its  action  does  not  extend  into  the 
surrounding  medium.  Plants  thus  prosper  by  the  help  of 
bacteria,  and  the  aim  of  cultivators  is  to  provide  for  each  plant 
the  microbes  most  favourable  for  the  yield  desired  by  mankind. 
But  it  is  to  be  noted  that  these  useful  microbes  are  not  within 
the  plant. 

The  problem  is  quite  otherwise  in  the  case  of  animals.  As 
in  so  many  other  cases,  the  question  was  first  stated  by  Pasteur 
in  connection  with  Duclaux's  experiments  on  the  nourishment 


MICROBES   IN  THE   HUMAN  BODY        29 

of  plants  under  sterile  conditions  and  their  rational  culture 
with  the  help  of  selected  microbes.  Pasteur  expressed  an 
opinion  which  he  had  not  time  to  submit  to  experiment,  but 
which  has  been  taken  up  by  others,  the  idea  namely  that  an 
animal  cannot  dispense  with  the  bacteria  living  within  it.  He 
suggested  that  a  young  animal  should  be  fed  from  birth  with 
pure  sterile  food. 

"  I  do  not  conceal  the  fact  that,  had  I  time  to  undertake 
this  study,  I  should  do  so  with  the  preconceived  opinion  that 
life  under  such  conditions  would  become  impossible.  Given 
that  such  experiments  are  capable  of  being  gradually  simplified, 
it  might  be  possible  to  study  digestion  by  adding  systematically 
to  the  sterile  food  of  which  I  speak  various  individual  bacteria 
or  different  bacteria  together,  each  of  definitely  known  species. 

"The  hen's  egg  lends  itself  without  serious  difficulty  to 
experiments  of  this  kind.  Previously  freed  externally  from 
every  sort  of  living  impurity  just  before  hatching,  the  chick 
should  be  immediately  put  into  a  chamber  free  from  every 
sort  of  bacterium,  so  that  it  could  be  supplied  with  pure  air 
and  with  sterile  food,  easily  introduced  from  without  in  the 
shape  of  water,  milk,  and  corn. 

"Whether  the  result  were  positive,  confirming  the  precon- 
ceived idea  which  I  am  putting  forward,  or  negative,  or  even 
absolutely  the  opposite,  t.e.,  that  life  without  bacteria  is  easier 
and  more  vigorous,  in  any  case  the  experiment  would  be  full 
of  interest." 

The  experiment  has  been  tried  and  the  programme  is  still  a 
long  way  short  of  completion.  One  thing  only  has  to  be 
altered  in  Pasteur's  statement.  The  experiments  are  far  from 
being  without  "serious  difficulty." 

Two  currents  of  opinion  have  developed.  The  first,  following 
literally  Pasteur's  idea,  maintains  that  animals  are  like  plants 
and  cannot  nourish  themselves  with  the  sole  help  of  their  own 
digestive  juices  but  require  also  the  intestinal  microbes.  We 
do  not  know,  say  its  supporters,  if  in  the  beginning  of  life  an 
animal  organism  has  ever  developed  free  from  micro-organisms ; 
it  is  not  very  probable ;  what  is  certain  is  that  for  centuries  and 


30  MICROBES   AND  TOXINS 

centuries  there  have  been  microbes  within  living  beings,  and 
there  must  have  become  established  a  sort  of  understanding,  or 
adaptation  between  the  intestinal  flora  and  the  intestine.  Just 
as  plant  roots  absorb  from  the  soil  the  fluids  elaborated  by 
the  bacteria,  so  the  intestinal  villi  absorb  the  juices  prepared 
by  the  microbes  of  the  intestine.  These  microbes  accumulate 
and  their  presence  in  masses  stimulates  the  intestinal  muscles. 
Further,  the  normal  intestinal  flora  opposes  an  invasion  by 
foreign  bacteria  which  might  be  or  might  become  pathogenic. 
Consequently  the  intestinal  bacteria  both  nourish  and  protect ; 
they  are  useful,  salutary,  and  providential. 

The  opposite  opinion  is  maintained  by  Metchnikoff.  The 
digestive  ferments  prepare  the  nutritive  materials  without  any  of 
this  problematical  assistance  from  the  microbes.  The  intestinal 
flora  is  injurious  (taking  of  course  the  point  of  view  which 
interests  us  most,  that  of  mankind),  because  long  before  the 
food  stuffs  leave  the  body,  the  bacteria  induce  in  them  fermenta- 
tions the  products  of  which,  absorbed  by  the  mucous  membrane, 
are  for  the  most  part  poisonous. 

From  the  point  of  view  of  nature,  it  is  quite  normal  for  the 
albuminous  excreta  of  our  food  to  putrefy  and  thus  return  into 
the  general  circulation  of  matter  ;  but  it  is  regrettable  when  this 
takes  place  in  our  bodies,  for  phenols,  skatol,  and  indol,  among 
other  products,  penetrate  into  our  circulation  and  affect  the  cells 
of  our  arteries  and  brain.  It  would  be  to  our  advantage  if  the 
food-stuffs  were  expelled  immediately  after  useful  digestion,  and 
before  the  terminal  phase,  the  putrefaction,  begins  in  the  waste 
products.  And  since  in  the  part  of  the  intestine  which  pro- 
perly speaking  is  the  digesting  part,  the  small  intestine,  there 
are  practically  no  bacteria,  and  since  on  the  other  hand  they 
swarm  in  the  large  intestine  where  there  are  scarcely  any  diges. 
tive  ferments,  it  is  evident  that,  on  the  whole,  the  intestinal 
flora  is  injurious.  The  ideal  condition  would  be  to  live  free 
from  bacteria  while  the  world  remained  populated  by  them. 
Is  a  life  of  such  purity  possible  ? 

Aseptic  breeding. — There  are,  it  has  been  said,  certain 
Arctic  animals,  both  birds  and  mammals,  without  intestinal 


MICROBES   IN   THE  HUMAN  BODY        31 

microbes.  But  the  recent  polar  expeditions  failed  to  find  a 
single  animal  of  this  kind. 

Nevertheless,  such  have  been  found  among  the  burrowing 
larvae  which  excavate  galleries  in  the  thickness  of  leaves  and 
live  shut  off  from  the  external  world  by  a  transparent  wall  of 
epidermal  cells.  The  truth  of  this  can  be  demonstrated  by 
extracting  them  with  a  sterile  needle  through  a  litttle  perforation 
made  in  the  epidermis  after  sterilisation  with  peroxide  of  hydro- 
gen. According  to  Portier's  experiments,  the  Lithocolktis 
caterpillars  are  aseptic  in  about  a  third  of  the  cases  ;  those  of 
the  Nepticula  of  rose-trees  always.  These  burrowing  larvae 
dispose  of  their  excreta  by  stowing  them  aseptically  in  their 
closed  tunnels,  whereas  Zischeria,  which  discharges  its  dejections 
externally,  through  a  little  hole  made  in  the  leaf,  is  always 
contaminated. 

Finding  experiments  on  these  small  invertebrates  easy, 
Bogdanoff  introduced  the  previously  disinfected  eggs  of  flies 
into  sterile  meat  and  found  that  the  larvae  hatched  and  kept 
under  these  conditions  develop  less  well  than  the  control  larvae 
fed  on  putrefying  meat  rich  in  bacteria.  When  he  added  to 
the  sterile  breeding  ground  a  digestive  ferment  capable  of 
attacking  meat,  i  e.y  trypsin,  the  sterile  larvae  flourished  equally 
with  the  others.  Bogdanoff  would  have  concluded  that  the 
bacteria  play  the  necessary  part  which  in  the  latter  case  was 
performed  by  the  ferment,  had  he  not  found  some  sterile  broods, 
without  addition  of  ferment,  quite  as  vigorous  as  in  ordinary 
breeding.  Larvae  can  thus  develop  w  ithout  the  aid  of  microbes. 
Wollmann  using  an  excellent  technique  has  finally  proved  this : 
he  succeeded  in  breeding  from  the  egg  sterile  flies.  "  During 
the  first  days  of  life  the  sterile  larvae  develop  more  slowly  than 
the  contaminated  controls,  probably  because  the  digestive 
glands  are  not  yet  in  full  activity  and  find  the  sterilised  meat 
difficult  to  attack.  Later  these  differences  disappear  and  the 
sterile  larvae  attain  the  weight  and  size  of  normal  adults." 

Experiments  on  aseptic  breeding  of  vertebrates,  such  as 
Pasteur  desired,  have  been  carried  out,  with  a  patience 
worthy  of  all  praise,  by  Nuttall  and  Thierfelder,  who  took  young 


32  MICROBES   AND  TOXINS 

guinea-pigs  from  their  mothers  by  Caesarean  section,  by 
Schottelius  and  Cohendy  on  the  chick,  by  Mme.  Metchnikoff 
and  by  Moro  on  tadpoles. 

The  little  guinea-pigs  appeared  to  develop  and  increase  in 
weight  in  normal  fashion,  but  Schottelius  has  questioned  the 
interpretation  of  these  experiments ;  he  maintains  that  t  he 
increase  in  weight  was  due  to  food  swallowed  but  not  digested. 
The  tadpoles  developed  badly  without  microbes.  The 
chickens  of  Schottelius  were  *  weaklings  '  and  lived  seventeen 
days  at  most,  becoming  increasingly  thin  and  feeble.  Their 
digestive  juices  were  apparently  insufficient  for  digestion,  and 
they  only  recovered  when  there  was  added  to  the  sterile  food 
certain  bacteria,  among  which  was  the  bacillus  coli. 

But  it  is  impossible  to  conclude  with  certainty  from  these 
experiments  that  life  is  impossible  without  microbes  ;  these 
new-born  animals  were  placed  under  conditions  too  different 
from  the  natural ;  their  intestine  was  not  yet  secreting  enough 
ferment,  and  the  food  they  were  getting,  sterilised  as  it  was  at 
a  high  temperature,  was  not  adapted  to  the  hereditary  disposi- 
tion of  their  alimentary  canal. 

It  would  therefore  be  necessary  to  study  adult  animals 
free  from  microbes,  but  we  know  that  this  condition  hardly 
exists.1 

There  are,  however,  animals  which  approach  it  somewhat, 
namely,  the  large  fruit-eating  bats  of  the  tropics  (Pteropus 
medius)  recently  studied  by  Metchnikoff.  These  animals 
have  a  large  intestine  very  limited  in  size ;  there  is  no 
reservoir  in  which  to  accumulate  the  waste  products  of  their 
food,  and  they  begin  to  evacuate  faeces  one  hour  after  the 
ingestion  of  the  food  from  which  these  are  derived.  They  are 
obliged  to  eat  a  great  deal  and  to  evacuate  in  proportion. 
They  have,  properly  speaking,  no  intestinal  flora,  simply  a 
few  bacteria  conveyed  by  the  food  and  changing  with  the  diet. 

1  The  intestine  of  the  scorpion  is  almost  always  free  from  microbes ; 
and  the  same  is  the  case  with  the  intestine  of  certain  maggots  provided 
with  digestive  juices  sufficiently  powerful  to  digest  seeds,  wool,  and  even 
such  resistant  microbes  as  the  bacillus  of  tubercle. 


MICROBES   IN   THE   HUMAN   BODY         33 

The  juices  of  their  small  intestine  have  no  power  of  killing 
bacteria  such  as  has  been  alleged  in  the  dog  and  the  cat,  so 
the  poverty  of  their  intestinal  flora  cannot  be  thus  explained. 
The  digestive  tube  being  almost  free  from  bacteria  in  its 
whole  length,  the  digestion  of  the  food  cannot  be  attributed  to 
microbes. 

The  fruit-bat  above-mentioned  digests  the  cellulose  of 
bananas,  yet  it  is  precisely  for  the  digestion  of  cellulose 
among  the  herbivora  that  the  necessity  of  auxiliary  microbes 
has  been  maintained  ;  the  reason  is  that  cellulose-digesting 
microbes  are  known,  whereas  no  ferment  has  been  separated 
from  the  digestive  tube  capable  of  doing  this.  But  the 
bacterial  ferment  has  not  been  isolated  either  :  the  question 
of  cellulose  digestion  is  one  on  which  our  ignorance  is  great 
and  which  demands  fresh  investigations.  Finally,  in  the 
dejecta  of  the  fruit-eating  bat  the  putrefactive  poisons  phenol, 
skatol,  and  indol  do  not  occur,  although  these  are  constant 
among  all  other  mammals  including  the  herbivora.  This 
absence  of  putrefaction  corresponds  with  the  extreme  short- 
ness of  the  large  intestine  and  the  poverty  of  the  intestinal 
flora.  We  have  then  in  this  animal  a  good  example  of  an 
adult  animal  capable  of  digesting  its  food,  under  normal 
conditions,  by  its  own  digestive  juices  without  the  help  of 
bacteria. 

The  Intestinal  Flora  of  Man. — Man  differs  from  the 
bat.  He  harbours  a  very  abundant  intestinal  flora  in  his  large 
intestine  which  is  highly  developed  and  in  which  the  food-stuffs 
begin  to  putrefy. 

It  is  almost  impossible  to  calculate  the  number  of  bacteria 
in  our  intestine.  Gilbert  and  Dominici  have  calculated  that 
we  discharge  daily  11,725  millions,  without  counting  the 
anaerobes,  of  which  there  is  a  prodigious  number.  Klein 
speaks  of  9  billions  of  bacteria  discharged  every  twenty-four 
hours,  of  which  100,000  millions  are  still  alive.  Strassburger, 
using  a  different  method  of  counting,  calculates  the  quantity 
of  bacteria  at  one-third  of  the  normal  dried  faeces  ;  in  twenty- 
four  hours  a  man  evacuates  120  billions  of  bacteria.  It  would 

D 


34  MICROBES   AND   TOXINS 

be  too  much  to  say  that  these  figures  are  imaginary,  but  they 
are  certainly  only  approximate.  It  is  sufficient  to  state  that 
the  real  figure  is  enormous  and  certainly  approaches  the 
magnitude  stated. 

The  small  intestine  is  less  populous,  so  much  so  that  a 
bactericidal  action  even  has  been  ascribed  to  it.  Yet  the 
pancreatic  juice  which  is  poured  into  the  small  intestine  is 
invaded  by  microbes  when  collected  and  left  outside  the  body. 
The  auto-sterilisation  of  the  small  intestine  seems  to  be  due  to 
the  combined  action  of  the  two  principal  digestive  juices,  the 
pancreatic  and  the  intestinal.  The  flora  of  the  small  intestine 
like  that  of  the  intestine  in  general  varies  with  the  diet. 

It  is  more  important  to  know  the  nature  of  the  microbes 
than  their  quantity.  In  1898  Mannaberg  counted  only 
twenty-seven  species  in  the  intestine  ;  but  there  are  now  many 
more  described  and  the  improvement  in  our  methods  of 
cultivation  is  leading  daily  to  the  discovery  of  new  ones. 
MacFadyean,  Nencki  and  Mme.  Sieber  have  isolated  fourteen 
from  the  small  intestine  alone. 

A  child  is  born  with  a  sterile  alimentary  canal,  but  from  the 
tenth  to  the  twentieth  hour  of  life  it  begins  to  be  inhabited. 


FIG.  9.  —  Bacillus  bifidus  (Tissier)  :  FIG.  10.  —  Bacillus  bifidus 

10  day  culture.  (Tissier)  :    15  day  culture. 

From  the  first  to  the  third  day  in  the  breast-fed  child  there 
appear  Streptococci,  Bacillus  coli,  B.  perfringens  (  =  Welch's 
Bacillus),  B.  Ill  of  Rodella,  B.  lactis  aerogenes,  Sarcinae, 


MICROBES   IN  THE   HUMAN  BODY        35 

enterococci,  mesentericus  and  an  acidophilus.  After  the  third 
day  the  flora  becomes  more  simple,  being  dominated  by  one 
of  the  last  to  appear,  the  anaerobe  B.  bifidus  of  Tissier,  which 
is  characteristic  of  the  flora  of  the  healthy  breast-fed  infant ; 
along  with  it  persists  the  Bacillus  coli,  the  enterococcus  and 
the  B.  lactis  aerogenes.  Thanks  to  the  presence  of  B.  bifidus 
and  the  lactic  bacilli,  which  produce  acids  from  the  carbo- 
hydrates (human  milk  is  rich  in  sugar  especially  after  the 
tenth  day),  the  putrefying  bacteria  (B.  perfringens,  for  example) 
are  kept  in  check  or  even  entirely  eliminated.  Proceeding 
from  the  stomach  to  the  rectum  the  species  which  predominate 
in  turn  are  the  B.  coli,  the  B.  lactis  aerogenes,  the  enUrococcus^ 


%  %        .*        *\  m 


FIG.  II. — Bacteria  in  the  FlG.  12. — Bacillus  sporogsnes 

feces  of  a  normal  child  of  (Metchnikoff). 

19  months. 

the  B.  exilis,  the  B.  acidophilus  and  lastly  the  B.  bifidus.  The 
distribution  and  proportions  may  be  altered  in  the  case  of  the 
infant  not  breast-fed.  The  chemical  surroundings  being 
different,  the  fermentations  also  differ. 

From  the  age  of  one  to  five  years  and  in  particular  after 
weaning,  the  flora  has  added  to  it  several  new  species,  while 
still  retaining  the  species  found  in  the  infant,  which  may  be 
regarded  as  the  fundamental  flora.  Tissier  calculates  the  pro- 
portion of  this  fundamental  flora  in  the  child  brought  up  on  a 
vegetable  diet  at  90  per  cent,  of  all  the  bacteria  and  J  of 

D    2 


36  MICROBES  AND  TOXINS 

this  are  made  up  of  B.  bifidus.  The  proportion  is  less  (70%) 
in  the  child  receiving  a  fair  quantity  of  animal  food. 

In  the  human  race  three  microbial  species  seem  to  be  chiefly 
capable  of  provoking  the  putrefaction  of  albuminous  material 
in  the  intestine ;  namely  the  three  anaerobes  :  B.  putrificus  of 
Bienstock,  B.  sporogenes  and  the  B.  of  Welch  also  called 
B.  perfringens. 

Not  only  are  these  three  anaerobes  putrefying  organisms  but 
they  seem  also  to  be  true  pathogenic  organisms.  Putrificus 
has  been  found  in  peritoneal  suppuration,  in  appendicitis  and  in 
various  intestinal  disorders.  B.  sporogenes  has  been  found  in 
many  cases  of  diarrhoea ;  B.  perfringens  is  common  in  acute 
and  chronic  suppurations,  and  in  infantile  diarrhoea  :  it  is  also 
the  cause  of  crepitating  gangrene  or  gaseous  phlegmon.  The 


FIG.  13. — Bacillus  FIG.  14. — Welch's  bacillus 

putrificus  (Bienstock).  (B.  perfringens). 

study  of  their  virulence  and  toxicity  by  experiments  on  small 
laboratory  animals  and  monkeys  has  hardly  been  begun.  They 
indicate  well  how  delicate  is  the  distinction  between  a  simple 
saprophyte  of  wide  prevalence  and  a  pathogenic  organism 
properly  speaking. 

The  B.  coli,  the  commonest  and  most  abundant  bacterium 
in  the  intestine,  though  it  was  considered  by  Schottelius,  in  his 
experiments  on  the  aseptic  chick,  as  an  indispensable  adjunct 
to  nutrition  and  though  it  has  even  been  employed  as  a  remedy 
for  constipation,  is  nevertheless  prejudicial  to  health. 

It  does  not,  it  is  true,  attack  the  genuine  albumins  but  it 


MICROBES   IN  THE   HUMAN   BODY        37 

breaks  down  the  peptones,  producing  indol,  phenol,  mercaptan 
and  sulphuretted  hydrogen.  It  is  precisely  these  bodies, 
putrefaction  products,  which  enter  the  blood  and  produce  the 
chronic  poisoning  which  induces  premature  old  age.1  Among 
the  bacteria  of  the  intestine,  B.  lactis  aerogenes,  B.  perfringens, 
B.  sporogenes^  Staphylococcus  pyogenes  and  Proteus  produce 
indol.  B.  coli  produces  both  indol  and  phenol,  and  there  is  ex- 
perimental proof  that  indol,  phenol,  and  their  sulpho-compounds 
can  produce  auto-intoxications  of  the  body.  With  indol  and 
potassium  phenyl-sulphate,  aortic  atheroma  has  been  produced 
in  rabbits  in  about  60  per  cent,  of  those  tried,  whereas 
spontaneous  atheroma  does  not  occur  in  more  than  6  to  10 
per  cent.  Monkeys  treated  with  paracresol  presented  arterial 
lesions  in  the  brain  and  kidney.  By  combining  the  phenols 
with  sulphur,  the  body  manufactures  "  sulpho-compounds ' 
which  are  less  injurious  than  the  pure  phenols,  but  which  are 
still  chronic  poisons  (MetchnikofT). 

Intestinal  Putrefaction. — The  study  of  the  putrefactive 
bacteria  being  in  its  infancy  and  beset  with  great  difficulties 
besides,  it  is  not  surprising  that  certain  authorities  maintain  that 
intestinal  putrefaction  is  harmless  to  man. 

It  is  true,  they  say,  that  our  instinct  leads  us  to  reject  putre- 
fying food  and  that  common  sense  has  always  connected 
putrefaction  with  disease.  Yet  we  see  the  Indo-Chinese,  the 
Malays,  Polynesians,  and  the  Greenlanders  regaling  them- 
selves with  decayed  fish,  meat,  or  eggs  !  Meat  which  has  gone 
bad  frequently  determines  diseases  resembling  acute  poisoning  ; 
but  it  is  not  because  it  is  putrefying  ;  it  is  because  it  contains 
pathogenic  bacteria  of  the  family  of  B.  typhosus,  or  bacilli  like 
the  B.  of  botulismus,  which  produces  a  violent  toxin.  Extracts 
of  putrefied  meat  have  been  injected  into  different  animals  with- 
out result.  Further,  how  many  people  exist  in  perfect  health  in 
spite  of  the  putrefaction  going  on  normally  in  their  intestine ! 
It  recalls  the  dictum  of  Malvoz  :  "  Tout  ce  qui  pue  ne  tue 

1  The  intestinal  flora  of  the  dog  does  not  differ  from  that  of  man. 
Little  rabbits  eight  days  old  have  not  a  rich  flora.  The  flora  of  parrots, 
young  and  old,  is  also  scanty  (five  species  in  the  ileum).  The  flora  of  the 
alligator  is  much  less  rich  than  that  of  man. 


38  MICROBES   AND   TOXINS 

pas,  tout  ce  qui  tue  ne  pue  pas."  (What  stinks  does  not  kill, 
what  kills  does  not  stink.) 

A  Belgian  physiologist,  Falloise,  has  shown  that  the  contents 
of  the  large  intestine  where  the  putrefications  are  going  on  are 
much  less  toxic  than  those  of  the  small  intestine  which  is  free 
from  putrefaction.  The  toxicity  of  faecal  matter  left  in  the 
incubator,  with  all  its  contained  bacteria,  diminishes  instead  of 
increases.  Hence  the  toxicity  is  not  in  proportion  to  the 
putrefaction. 

What  then  causes  those  gastro-intestinal  disorders,  in 
particular  infantile  diarrhoea,  with  symptoms  resembling  typhoid 
fever  or  cholera  ?  It  must  be  poisons  of  some  kind,  but  poisons 
which  are  the  result  not  of  bacterial  decomposition  of  the 
intestinal  contents  but  of  abnormal  metabolism  of  the  food. 

This  theory  of  non-microbial  intoxication,  is  opposed  to  that 
which  regards  microbial  putrefaction  as  the  cause  of  the  poisoning. 
Finkelstein,  for  example,  says  that  there  is  no  specific  infecting 
organism  in  infantile  diarrhoea  and  that  the  same  symptoms 
appear  with  very  different  intestinal  flora  ;  according  to 
Nobecourt  and  Rivet,  the  intestinal  flora  depends  on  the  diet 
and  the  disorders  of  digestion,  its  role  being  secondary  and 
rather  an  effect  than  a  cause.  The  principal  role  is  played  by 
those  poisons  which  are  produced  directly  in  the  digestion  of 
the  food-stuffs. 

But  though  it  is  true  that  the  study  of  intestinal  putrefaction 
is  not  far  advanced,  the  opponents  of  this  theory  must  acknow- 
ledge that  still  less  is  known  about  the  auto-intoxications.  Nothing 
definite  has  been  established  in  connection  with  the  ptomaines 
and  toxalbumins  which  have  been  incriminated.  Besides,  the 
argument  of  Falloise  on  the  slight  toxicity  of  the  faecal  matter 
is  not  justified,  for  he  forgets  that  the  faeces  do  not  represent 
actually  the  intestinal  contents,  these  latter  having  lost 
before  discharge  precisely  those  poisons  which  have  been 
absorbed  by  the  mucous  membrane. 

The  phenols,  for  example,  which  are  produced  in  our  intestine 
are  absent  in  the  faeces,  but  appear  in  the  urine.  The  volatile 
fatty  acids  also  are  in  greater  quantity  in  the  urine  than  in  the 


MICROBES   IN  THE   HUMAN   BODY        39 

faecal  matter.  As  to  the  poisons  found  in  the  small  intestine, 
these  are  nothing  but  the  digestive  juices  themselves,  from  the 
pancreas  and  intestinal  mucous  membrane ;  the  fact  of  their 
toxicity  when  injected  into  the  veins  of  a  rabbit  does  not  prove 
in  the  least  that  they  are  noxious  to  the  animal  producing 
them.  It  is,  in  fact,  a  very  rough  way  of  studying  the  toxicity 
of  faecal  matter  to  inoculate  extracts  of  it  into  animals.  The 
biological  and  chemical  study  of  the  intestinal  bacteria  in  their 
capacity  as  ferments  is  the  necessary  and  proper  introduction  to 
the  problem  of  intestinal  putrefaction. 

The  diminution  of  toxicity  which  Falloise  discovered  on 
incubating  faeces  represents  only  the  primary  phase  of  the 
phenomena  which  occur  under  such  conditions,  namely,  a 
preliminary  fermentation  which  produces  acid  and  suspends  the 
putrefaction  ;  it  is  only  after  the  third  day  that  true  putrefaction 
can  begin,  the  reaction  having  then  become  alkaline. 

Finkelstein's  argument  was  the  speedy  effects  of  change  of 
diet  on  an  intestinal  disorder  ;  but  this  action  may  equally  well 
be  explained  by  the  intervention  of  microbes ;  the  flora  also 
varies  with  the  diet.  In  the  intestine,  as  in  a  culture  flask, 
different  cultures  can  be  got  with  the  same  microbes  when  the 
media  originate  on  the  one  hand  from  meat  and  on  the  other 
from  vegetables. 

In  studies  so  difficult  as  these,  no  good  result  can  be  got  by 
treating  faecal  material  and  injecting  it  in  quantity.  It  is 
necessary  to  proceed  analytically,  isolating  patiently  the 
bacterial  species  of  the  intestines,  studying  them  in  a  condition 
of  purity,  as  every  ferment  ought  to  be  studied,  studying 
secondly  their  actions  in  association,  and  studying  finally  the 
poisons  they  produce,  phenols,  indol,  skatol,  etc.  It  is  by 
this  method  that  the  action  of  B.  putrificus,  B.  perfringens, 
and  B.  sporogenes  in  intestinal  decomposition  has  been 
rendered  certain  as  has  been  the  role  of  B.  proteus  in  infantile 
diarrhoea. 

Mutual  Assistance  and  Mutual  Antagonism. — These 
legions  of  microbes  do  not  escape  the  law  of  competition  which 
prevails  throughout  life.  They  have  not  all  the  same  demands 


40  MICROBES   AND  TOXINS 

or  the  same  appetites ;  one  requires  more  alkalinity,  another 
requires  a  certain  degree  of  acidity.  The  unused  residue  of  the 
food  of  one  may  provide  excellent  nourishment  for  another. 
Their  actions  on  elementary  waste  products  have  to  follow  each 
other  in  a  regular  and  chemically  definite  order.  Hence  we  get 
agreements  and  antagonisms,  one  species,  by  reason  of  its 
nature  and  the  food  of  its  host,  being  towards  another  either 
a  help  or  a  hindrance. 

This  idea  was  first  expressed  in  connection  with  the  bacteria 
of  the  intestine  by  the  experiments  of  Metchnikoff  on  cholera 
and  its  vibrio.  Cholera  is  due  to  the  multiplication  in  the 
intestine  of  a  spirillum,  which  rarely  penetrates  into  the  blood, 
but  elaborates  in  the  intestine  poisons  which  diffuse  and  kill 
the  patient.  Cholera  is  an  acute,  toxic,  specific  enteritis. 
Koch's  discovery  of  the  spirillum  or  "  comma  bacillus "  of 
cholera  had  to  contend  with  an  obstinate  scepticism,  because 
cholera  could  not  be  produced  with  it  at  will  in  laboratory 
animals,  unlike  anthrax  or  fowl-cholera.  Whether  by  inoculation 
under  the  skin,  or  in  the  peritoneum,  or  by  the  mouth,  the 
result  was  the  same.  Even  when  several  savants  swallowed 
cultures  of  it,  the  results  of  these  experiments  "in  anima  nobili" 
were  very  inconstant.  In  this,  typhoid  fever  resembles  cholera, 
the  disease  being  very  difficult  to  reproduce  experimentally; 
it  is  only  quite  recently  that  experimental  typhoid  fever  has 
been  successfully  produced  by  employing  the  higher  apes,  the 
nearest  congeners  of  man. 

It  cannot  be  a  matter  of  indifference  to  the  cholera  vibrio 
what  is  the  nature  of  the  flora  of  the  alimentary  canal  into 
which  it  penetrates.  By  a  very  simple  method  of  experiment, 
namely,  by  inoculating  together  on  gelatine  plates  vibrios  and 
various  other  microbes  along  lines  which  crossed  each  other, 
Metchnikoff  was  able  to  prove  that  the  association  of  certain 
bacteria  favoured  the  growth  of  cholera  while  others  inhibited 
it.  Round  the  colonies  of  a  favouring  bacterium  a  swarm 
of  little  cholera  colonies  developed.  Various  favouring  and 
inhibiting  bacteria  were  discovered  in  the  air,  among  animals, 
and — most  interesting  of  all — in  the  human  stomach. 


MICROBES   IN   THE   HUMAN   BODY         41 

But  here  we  were  still  only  dealing  with  growth  on  plates, 
i.e.,  under  very  artificial  conditions.  The  importance  of  the  ex- 
periments increased  when  it  was  found  possible  to  transmit  to 
an  animal,  with  certainty,  a  genuine  attack  of  cholera.  Neither 
the  cat  nor  the  guinea-pig  nor  the  adult  rabbit  takes  the 
disease.  Metchnikoff  had  the  idea  that  their  resistance  might 
be  due  to  the  presence  in  the  alimentary  canal  of  inhibiting 
bacteria,  and  he  experimented  accordingly  on  little  sucking 
rabbits,  which,  while  they  are  at  the  breast — a  period  of  some 
weeks — present  a  very  scanty  intestinal  flora.  Little  rabbits  of 
one  to  four  days  old,  made  to  swallow  a  culture  of  the  cholera 
vibrio,  died  in  about  half  the  cases  ;  they  were  thus  capable  of 
taking  cholera.  But  further,  when  to  the  culture  of  the  vibrio 
other  bacteria  were  added  which  had  been  recognised  as 
being  favouring  in  the  gelatine  experiment,  the  little  rabbits 
took  cholera  without  exception.  They  were  refractory,  on  the 
contrary,  when  the  inhibiting  bacteria  were  added. 

The  question  naturally  arose  whether  during  an  epidemic 
man  might  not  increase  his  resistance  to  a  cholera  infection 
by  ingesting  cultures  of  these  inhibiting  microbes,  and  this 
formed  the  starting  point  of  intestinal  bacteriotherapy. 

Bienstock  attributed  the  resistance  which  milk  possesses 
naturally  towards  putrefaction  to  the  microbes  which  it  contains, 
and  not,  as  the  old  idea  maintained,  to  the  presence  in  it  of 
casein  or  milk  sugar ;  he  found  that  the  B.  lactis  aerogenes,  an 
acid  producer,  and  even  the  B.  coli,  prevent  the  development 
of  the  B.  putrificus,  the  agent  of  putrefaction,  whether  in  an 
experimental  flask  or  in  the  intestine.  Inoculated  along  with 
the  B.  coli,  the  growth  of  B.  putrificus  is  inhibited,  because  it 
can  only  develop  in  an  alkaline  medium,  and  the  B.  coli 
produces  from  the  sugars  of  the  food  materials  an  abundance 
of  acid. 

The  idea  of  an  inhibiting  action  exerted  by  an  acid-producing 
bacterium  on  the  putrefying  organisms  is  correct,  but  the  above 
example  was  not  the  best ;  in  the  first  place  because  the 
B.  putrificus  is  rare  in  the  human  intestine,  and  secondly 
because  the  B.  coli,  while  playing  a  useful  part  in  certain  cases, 


42  MICROBES  AND  TOXINS 

is  in  others  itself  a  putrefying  organism.  Kolbriigge's  opinion 
can  no  longer  be  maintained  that  the  B.  colt  is  the  beneficent 
bacterium  par  excellence,  growing  in  the  appendix  as  in  a  sort  of 
hot-house  put  specially  there  by  Providence ! 

The  antagonism  is  chiefly  between  the  B.  perfringens  of 
Welch  and  the  B.  bifidus  discovered  by  Tissier  in  the  normal 
flora  of  the  healthy  breast-fed  infant. 

There  is  a  general  antagonism  between  the  simple  saccharo- 
lytic  bacteria  which  produce  acid  from  starches  and  sugars 
and  the  simple  proteolytic  bacteria  which  carry  the  digestion  of 
meat  to  the  stage  of  indol,  skatol,  phenol,  and  ammoniacal 
salts.  When  the  bacteria  which  oppose  each  other  are  both 
proteolytic  and  saccharolytic  (hence  called  mixed  ferments), 
those  which  produce  the  most  acid  inhibit  those  which  produce 
less. 

For  example,  the  enterococcus  is  inhibited  by  an  acidity  of 
2 '45  per  1000;  the  B.  perfringens  at  r6o  per  1000 ;  the  B. 
acidi  paralactid  at  5*39;  the  B.  bifidus  at  4-90.  The  two 
last  are  evidently  successful  antagonists  of  the  B.  perfringens. 

The  facts  established  by  Bienstock  and  by  Tissier  and 
Martelly  may  be  expressed  as  the  following  law :  in  media 
containing  protein  along  with  more  than  10  per  1000  of  sugar, 
a  "  mixed  ferment "  can  arrest  the  development  and  the 
action  of  a  simple  ferment,  whereas  a  strong  mixed  ferment 
(i.e.,  a  strong  acid  producer)  can  inhibit  a  weaker.  These 
inhibiting  actions  are  entirely  due  to  the  quantity  of  acid 
which  the  bacteria  produce  in  the  course  of  their  consumption 
of  the  carbohydrates. 

Principles  of  Intestinal  Bacteriotherapy.— 
Metchnikoffs  experiments  on  microbial  associations  in  cholera 
contain  the  germ  of  a  method  of  treatment  which  consists  in 
using  inhibiting  bacteria.  The  experiments  of  Bienstock 
and  Tissier  indicate  that  the  saccharolytic  bacteria  form  a 
natural  obstacle  to  the  action  of  the  putrefying  microbes. 
The  idea  of  a  bacteriotherapy  had  already  been  formulated  by 
the  well-known  children's  physician,  Escherich,  practically  in 
the  following  terms :  "  It  is  necessary  to  employ  the  '  acid 


MICROBES   IN   THE   HUMAN    BODY        43 

fermentations  *  as  antagonists  of  the  '  alkaline  fermentations, 
either  by  adding  to  the  diet  carbohydrates  such  as  lactose  (milk 
sugar),  or  by  giving  cultures  of  acid-producing  bacteria." 

It  had  long  been  the  custom  to  employ  as  remedies  for 
diarrhoea,  intoxications,  and  intestinal  putrefactions,  first 
purgatives,  then  antiseptics,  of  which  the  most  popular  was 
/3-naphthol.  This  treatment  has  been  given  up. 

Several  days  of  /?-naphthol  treatment  fail  to  diminish  the 
number  of  bacteria  in  the  alimentary  canal  (Stern). 

"  Since  intestinal  antisepsis  is  based  on  such  a  weak  found- 
ation," said  Quincke  in  a  treatise  on  intestinal  auto-intoxi- 
cations, "it  was  natural  to  search  for  another  method  of 
combating  the  decomposition  caused  by  bacteria  in  the 
intestine,  and  the  idea  came  to  me  to  try  to  supplant  the 
injurious  bacteria  by  beneficent  ones,  just  as  weeds  are 
suppressed  by  the  growth  of  useful  crops,  the  starting  point 
being  the  antagonism  of  different  microbes  observed  in 
artificial  culture  media."  Quincke  tried  the  yeast  of  beer. 
Later  there  was  employed  in  infantile  diarrhoea  the  Bacterium 
lactis  aerogenes,  the  antagonist  of  B.  proteus. 

Intestinal  bacteriotherapy  has  now  become  part  of  practical 
medicine  and  is  applied  in  two  principal  forms.  The  first  is 
to  administer  those  foods  which  are  produced  by  natural 
fermentations  and  which  contain  great  numbers  of  bacteria; 
the  second  is  to  give  along  with  a  suitable  diet  artificial  cultures 
of  bacteria  possessing  antiputrefactive  properties.  In  both 
cases  it  is  the  antagonistic  action  of  microbes  towards  the 
bacteria  of  decomposition  of  which  one  takes  advantage. 

Fermented  foods,  rich  in  bacteria,  were  employed  long 
before  modern  researches  on  microbial  antagonism  came  into 
being. 

Humanity  has  known  from  time  immemorial  the  whey  of 
milk  and  the  various  forms  of  cheese.  It  was  long  before 
the  days  of  science  that  there  were  invented  koumiss,  kvass, 
kephir,  lebenraib  and  yoghourt.  In  the  Bible  we  read  that 
Abraham  offered  to  his  guests  sour  milk  as  well  as  sweet 
Moses  says  that  Jehovah  has  given  his  people  to  eat  of  the  sour 


44  MICROBES   AND   TOXINS 

milk  of  cows  and  the  milk  of  goats  (at  the  same  time,  it  is  true, 
the  fat  of  lambs  and  of  sheep  and  of  kidney).  There  can  be 
no  doubt  that  sour  milk  is  more  ancient  even  than  the  Bible 
itself. 

The  artificial  cultures  chosen  for  their  chemical  properties 
are  chiefly  B.  bifidus,  B.  acidi paralactici  and  the  lactic  ferment 
which  has  become  popular  under  the  name  of  the  Bulgarian 
bacillus ;  they  are  given  either  singly  or  in  association.  They 
may  be  taken  either  in  the  form  of  clotted  mik  or  in  bouillon. 
The  characters  of  the  Bulgarian  bacillus  are  briefly  the 
following  :  it  produces  25  grams  of  lactic  acid  per  litre  of  milk; 
not  more  than  0*50  gram  of  succinic  and  acetic  acid,  traces  of 
formic  acid,  no  alcohol,  and  no  acetone.  It  hardly  attacks  the 
proteins  at  all,  and  has  no  pathogenic  power  (G.  Bertrand  and 
Weisweiler).  Although  it  does  not  inhibit  the  development  of 
the  B.  coli  in  a  culture  containing  peptone,  it  at  least  prevents 
it  from  producing  phenol  and  greatly  reduces  its  indol 
production. 

The  mechanism  of  the  action  of  the  lactic  ferments  in 
general  is  not  quite  settled.  They  appear  to  diminish  the 
number  of  anaerobes  and  of  the  B.  coli.  A  diet  of  sour  milk 
reduces  the  ethereal  sulphates  of  the  urine  more  than  does  a 
diet  of  sweet  milk. 

The  Large  Intestine  is  a  Useless  Organ. — Since  the 
large  intestine  performs  no  digestive  action  and  absorbs  poison, 
it  follows  that  it  is  not  only  a  useless  but  an  injurious  organ. 
Medicine  may  possibly  perfect  methods  of  treatment  and  diets 
to  prevent  it  from  doing  harm  ;  surgery  perhaps  will  reach  this 
end  more  quickly.  However  that  may  be,  certainly  no  one 
has  ever  proposed  that  the  large  intestine  should  be  removed 
from  every  human  being ;  but  it  is  not  the  less  true  that  from 
the  zoological  point  of  view  it  is  a  useless  inheritance  left  to  us 
by  our  animal  ancestors.1 

1  We  must  consider  it  as  a  useless  inheritance  from  our  ancestors  during 
evolution,  though  they  no  doubt  derived  some  benefit  from  it.  Compara- 
tive anatomy  teaches  us  that,  among  all  the  vertebrates,  it  is  only  the 
mammals  which  are  provided  with  a  large  intestine  properly  speaking. 
Birds,  reptiles,  and  other  lower  vertebrates  do  not  possess  one 


MICROBES   IN  THE   HUMAN  BODY        45 

Examples  of  human  beings  who  have  lived  after  the  partial 
or  total  removal  of  the  large  intestine  or  after  the  suppression 
of  its  function  are  far  from  being  rare.  A  patient  of  Korte 
survived  eight  operations  on  his  intestine  ;  another  of  Wiesinger 
remained  in  a  satisfactory  condition  after  losing  both  his  trans- 
verse and  descending  colon.  Another  of  Ciechomski  lived  for 
thirty  years  with  a  fistula  which  had  put  the  large  intestine  out 
of  use ;  he  married  and  had  children.  Tavel's  patient  (studied 
by  MacFadyean,  Nencki,  and  Sieber),  aged  sixty-two,  had  to 
digest  her  food  for  six  months  without  the  large  intestine,  yet 
she  remained  well.  A  patient  of  Mauclaire  has  been  living 
for  years  in  fair  health  with  her  large  intestine  out  of  use. 
The  histories  of  these  patients  are  to  be  found  in  the  most 
respectable  surgical  journals. 

An  English  surgeon,  Lane,  recently  published  an  account  of 
thirty-nine  removals  of  the  large  intestine,  partial  or  complete  ; 
he  lost  nine  out  of  the  number,  i.e.,  23  per  cent.  Many  of  the 
survivors,  formerly  subject  to  enteritis  and  severe  neurasthenia, 
had  never  been  able  to  enjoy  life  till  it  had  become  thus 
"  simple."  The  mortality  is  still  too  great  for  this  operation 
to  be  anything  but  an  exceptional  procedure.  But  surgical 
technique  is  progressing  every  day.  What  surgeon  would 

It  is  probable  that  the  large  intestine  acted  as  a  reservoir  for  the  waste 
products  of  digestion.  The  mammals  having  to  run  fast  to  escape  from 
their  enemies  or  to  capture  their  prey,  were  inconvenienced  by  the 
necessity  of  stopping  to  empty  their  bowels.  A  large  intestine  under 
these  conditions  would  be  of  the  greatest  use.  Thus  we  see  that  those 
mammals  which  run  fastest,  such  as  the  horse  and  the  hare,  possess  the 
most  highly-developed  large  intestine  and  caecum.  It  is  remarkable  that, 
among  birds,  the  runners  such  as  the  ostriches  and  cassowaries  have 
similarly  acquired  a  large  intestine  and  possess  cseca  more  developed  than 
all  the  other  feathered  creatures.  It  is  thus  the  demand  made  by  the 
struggle  for  life  which  has  led  to  the  formation  of  the  large  intestine 
among  the  vertebrates.  The  high  development  of  this  organ  as  a 
reservoir  for  the  solid  excreta  has  produced  in  its  turn  the  development  of  a 

very  rich  bacterial  flora Man  having  no   necessity  for  the   large 

intestine  and  its  flora  either  for  the  digestion  of  cellulose  or  for  prolonged 
retention  of  the  residues  of  digestion  derives  no  advantage  whatever  from  its 
possession.  On  the  contrary  he  suffers  very  numerous  inconveniences. 
It  is  easy  to  conceive  how  an  organ  thus  become  useless  con- 
tributes to  the  shortening  of  life."  Metchnikoff,  Wilde  Lecture,  Manchester 
Memoirs,  1901. 


46  MICROBES   AND  TOXINS 

have  prophesied  fifty  years  ago  that  the  removal  of  the 
appendix  would  become  as  simple  an  operation  as  the 
extraction  of  a  tooth  ? 

From  the  scientific  point  of  view  it  is  quite  legitimate  to 
conclude  that  in  a  mammal  so  advanced  in  evolution  as  is 
civilised  man,  both  life  and  digestion  are  possible  in  the  absence 
of  a  large  intestine. 

Is  the  Intestine  Permeable  for  Microbes  ? — There 
exist  in  the  massive  flora  of  the  intestine,  bacteria  both  actually 
and  potentially  pathogenic.  After  a  wound  which  penetrates 
the  intestinal  wall,  the  bacteria,  which  were  harmless  while  in 
the  alimentary  canal,  invade  the  general  peritoneum  and  cause 
a  fatal  peritonitis.  Immediately  after  death  the  intestinal 
bacteria  invade  all  the  tissues  and  begin  the  work  of  putre- 
faction. To  prevent  this  happening  during  the  lifetime  of  the 
animal  it  is  obviously  necessary  for  the  intestine  to  be 
impermeable,  and  this  impermeability  must  be  due  to  the 
living  healthy  mucous  membrane. 

This  opposing  force  is  sometimes  thrown  out  of  action. 
There  are  certain  diseases  which  can  only  be  explained  by 
supposing  that  the  pathogenic  bacterium  has  passed  from  the 
intestine  into  the  blood  and  into  the  organs.  In  hog-cholera, 
which  is  caused  by  a  virus  still  unknown,  the  lesions  of  the 
lung  contain  a  microbe  which  develops  under  cover  of  the 
true  virus  and  forms  a  regular  complication  in  this  disease. 
This  microbe  is  a  normal  inhabitant  of  the  intestine  of  healthy 
pigs.  In  all  infectious  diseases  in  which  infection  occurs  by 
ingestion,  and  which  are  not  exclusively  confined  to  the 
alimentary  canal,  it  is  necessary  to  suppose  that  the  microbe 
passes  from  the  intestine  into  the  blood  either  directly  by  the 
capillary  vessels  of  the  mucous  membrane  or  by  the  more 
roundabout  way  of  the  mesenteric  lymphatic  glands.  Typhoid 
fever,  for  example,  is  mainly  a  disease  of  the  intestine,  an 
enteritis,  but  during  the  whole  course  of  the  fever  the  bacillus 
circulates  in  the  blood,  and  occasionally  forms  colonies  in 
distant  organs  such  as  the  lungs  and  the  bones. 

A  very  simple   explanation   suggests   itself:   the  intestinal 


MICROBES  IN  THE   HUMAN   BODY        47 

barrier  has  been  pierced  by  a  little  wound  permitting  the 
passage  of  the  bacteria.  This  happened  in  the  case  of  Pasteur's 
sheep,  which  took  anthrax  when  he  fed  them  with  anthrax 
spores  mixed  with  minute  splinters.  But  this  explanation  does 
not  hold  for  all  cases.  Numerous  facts  exist  which  establish 
the  permeability  of  the  healthy  mucous  membrane. 

The  importance  of  the  problem  was  perceived  when  the 
conditions  of  infection  with  tubercle  began  to  be  discussed. 
Behring  thought,  in  agreement  with  Chauveau,  that  the  bacillus 
penetrates  into  the  body  of  man  and  cattle,  not  only  by  the 
respiratory  passages,  but  also  through  the  digestive  tract.  This 
idea  was  suggested  to  him  by  the  predominance  of  mesenteric 
lesions  in  infantile  tuberculosis,  compared  with  the  predomin- 
ance of  pulmonary  lesions  in  the  adult;  he  thought  that 
tuberculosis  always  began  in  childhood  in  the  intestine  and 
did  not  invade  the  lung  until  maturity.  The  intestine  of  an 
adult,  he  maintained,  does  not  permit  the  passage  of  the 
bacteria  as  does  that  of  a  child,  the  reason  being  that  in  the 
infant,  as  in  general  amang  new  born  animals,  the  intestinal 
mucosa  is  not  coated  with  the  continuous  and  perfect  covering 
which  develops  later ;  the  covering  is  discontinuous,  inter- 
rupted and  open  to  the  passage  of  bacteria.  No  intestinal 
lesion  can  be  found  because  none  exists :  there  has  been 
penetration  without  violence. 

Behring's  idea  has  been  developed  to  such  an  extent  that 
some  observers  declare  that  invasion  by  the  intestine  is  more 
frequent  than  invasion  by  the  lung  ;  and  during  the  search  for 
proofs  and  comparisons  they  have  studied  closely  the  penetra- 
tion of  the  intestine  not  only  by  the  bacillus  of  tuberculosis, 
but  by  various  other  bacteria,  pathogenic  and  non-pathogenic, 
and  even  by  inert  dust  particles.  Pulmonary  anthracosis,  the 
disease  or  rather  the  histological  condition  which  consists  in  the 
impregnation  of  the  lungs  by  carbon  particles  from  the  air  of 
mines  and  factories,  has  formed  the  field  of  battle  for  the 
partisans  and  opponents  of  Behring's  idea,  and  a  multitude  of 
experiments  have  resulted,  not  without  instructive  consequences. 

Dust  particles  can  reach  the  lung  by  passing  through  the 


48  MICROBES    AND   TOXINS 

intestine,  but  that  is  not  the  rule  and  can  only  be  produced  by 
administering  the  dust  in  large  quantities  in  rapid  succession  : 
these  are  artificial  and  exceptional,  not  natural,  conditions.  It 
is  the  same  in  the  case  of  bacteria.  In  an  animal  in  perfect 
health  they  do  not  penetrate  the  intestinal  wall,  but  they  can 
be  made  to  do  so  under  the  influence  of  fasting  and  fatigue 
(experiments  of  Ficker).  One  may  say  that  their  passage  is 
the  first  and  a  very  early  sign  of  an  agonal  phenomenon,  an 
anticipation  of  that  exodus  of  bacteria  which  occurs  immed- 
iately after  death. 

We  must  not  forget,  however,  that,  according  to  the  state- 
ments of  Porcher  and  Desoubry,  bacteria  pass  from  the 
intestine  into  the  blood  each  time  the  animal  digests,  and  that 
to  get  sterile  sera  it  is  necessary  to  bleed  anti-diphtheria  and 
anti-tetanus  horses  only  during  the  intervals  between  digestion. 

The  question  is  less  simple  than  it  looks,  because  the  mucous 
membrane  is  quite  as  active  as,  if  not  more  so  than,  the 
bacteria.  Not  only  is  it  built  up  of  active  living  cells,  but  there 
are  continually  traversing  it  other  living  motile  cells,  the 
leucocytes,  which  play  an  active  part  in  the  phenomena  of 
digestion  and  absorption,  of  defence  and  of  resistance. 

The  cellular  mechanism  of  defence  does  not  invalidate  the 
conclusion  of  this  chapter.  Being  useless  for  digestion,  and  a 
constant  cause  of  putrefaction,  the  intestinal  flora  is  a  perman- 
ent source  of  danger  for  the  body.  The  intestinal  flora 
represents  in  our  interior  the  external  world  with  its  unregulated 
fermentations  only  limited  by  their  mutual  competition.  It 
represents  in  the  interior  of  our  bodies  the  innumerable  legions 
of  bacteria  in  nature,  which  find  their  pabulum  and  their  prey 
in  fermentable  material  of  every  kind,  and  which  are  as  eager 
to  attack  the  proteins  and  carbohydrates  of  our  living  tissues 
as  to  attack  the  sugar  and  the  casein  of  fermenting  milk.  The 
power  of  resistance  of  the  body  is  counterbalanced  by  its 
sensitiveness  towards  the  poisons  which  are  being  continually 
secreted,  and  which  are  capable  at  least  of  accelerating  the 
approach  of  old  age  if  not  of  directly  causing  death. 

Researches   on  the  intestinal   flora   are   bound   to   take   a 


MICROBES  IN  THE   HUMAN   BODY        49 

greater  and  greater  place  in  experimental  medicine.  With  the 
help  of  their  results  microbiology  will  then  attack  the  problem 
of  those  chronic  diseases,  diseases  of  nutrition,  such  as  gout, 
rheumatism,  and  diabetes.  .Finally  the  discovery  of  the  so- 
called  invisible  viruses  proves  that  the  field  of  action  of  bacteria 
is  much  wider  than  had  been  suspected  and  that  there  may 
well  exist  legions  of  active  bacteria  in  situations  where  none 
have  yet  been  seen. 


CHAPTER   III 

FORM    AND    STRUCTURE   OF    MICROBES 

Animal  and  vegetable  microbes — Protozoa— Moulds  and  bacteria. 
Morphology  of  bacteria — Membrane  ;  vibratile  cilia,  capsules,  Multiplica- 
tion, Spores. 

Pleomorphism  of  bacteria — The  place  of  bacteria  in  a  general  classification. 
The  nucleus  in  bacteria  :  the  diffuse  nucleus. 
Reproduction  and  the  possession  of  sex  among  microbes. 
Chemical  composition. 

"  MICROBES  "  is  the  name  given  to  those  living  beings  which 
can  only  be  seen  by  the  aid  of  the  microscope.  They  are  the 
simplest  organisms  found  in  nature.  Some  belong  to  the 
animal  kingdom,  others  to  the  vegetable.  The  bacillus  of 
tuberculosis  and  the  typhoid  bacillus  belong  to  the  class  of 
bacteria,  and  the  bacteria  are  plants.  The  trypanosoma  of 
sleeping  sickness  is  an  infusorian,  i.e.,  an  animal. 

Whether  animal  or  vegetable,  almost  all  microbes  consist  of 
a  single  cell.  When  there  are  several  cells,  they  are  practically 
identical  individual  cells  in  juxtaposition. 

The  tissues  and  organs  are  systems  of  cells ;  microbes 
possess  neither  tissues  nor  organs. 

Like  every  cell  a  microbe  possesses  a  nucleus  and  proto- 
plasm. 

The  vegetable  cell  is  enveloped  by  a  membrane  whose 
composition  is  not  well  known,  and  even  the  motile  ones  are 
more  rigid  than  the  animal  microbes. 

Microbes  are  of  all  living  creatures  the  most  widespread. 
They  exist  wherever  there  is  organic  matter. 

Protozoa. — In   a   hay  infusion — a   handful  of   hay  in  a 

50 


FORM  AND  STRUCTURE  OF  MICROBES     51 

vessel  of  water — one  can  see  under  the  microscope  motile 
creatures,  whose  surface  is  furnished  with  cilia,  whiplets  or 
flagella  in  vibration  ;  these  are  the  infusoria,  some  with  flagella, 
others  with  cilia.  Sleeping  sickness  is  caused  by  a  trypanosoma^ 
a  flagellate  protozoon  found  at  certain  times  in  the  blood, 
in  the  lymphatic  glands  of  the  neck  and  in  the  cerebro-spinal 
fluid.  These  protozoa  resemble  minute  flattened  worms  in 
the  interior  of  which  there  are  two  nuclei,  one  large,  one 
small ;  on  one  side  there  is  an  undulating  membrane  bordered 
by  the  flagellum,  which  is  continued  beyond  the  anterior  end. 


FIG.  15.— -Trypanosomes  and  blood  corpuscles. 

The  trypanosomes  multiply  by  longitudinal  division  beginning 
with  the  nucleolus  and  continuing  through  the  flagellum,  the 
large  nucleus,  and  the  protoplasm.  There  is  a  possibility  that 
sexual  reproduction  exists. 

There  is  often  found  in  the  intestine  of  rabbits  an  animal 
parasite  which  passes  through  a  complicated  series  of  different 
forms ;  it  possesses  a  cycle  of  evolution,  one  portion  in  the 
rabbit,  one  portion  outside.  This  parasite  is  a  coccidium 
belonging  to  the  class  of  sporozoa.  The  microbe  of  malaria 
is  a  parasite  of  the  blood  corpuscles,  a  haematozoon,  which  has 
for  long  been  regarded  as  related  to  the  coccidia,  but  which 

£  2 


52  MICROBES  AND  TOXINS 

possesses  certain  characters,  inclining  one  rather  to  derive  it  from 
the  flagellates.  It  too  passes  through  a  cycle  of  development, 
one  stage  being  in  the  blood  of  man,  the  other  in  the  body  of 
a  mosquito  of  the  genus  Anopheles.  According  to  the  stage 
of  development,  the  hsematozoa  of  Laveran  have  both  asexual 
and  sexual  reproduction,  the  latter  occurring  in  the  body  of 
the  mosquito. 

It  cannot  be  too  often  insisted  on  that  no  protozoon  is 
properly  known  until  all  its  cycle  of  development  has  been 
investigated  and  settled. 

Moulds. — A  slice  of  bread  or  a  piece  of  lemon  left  to 
itself  under  suitable  conditions  of  temperature  and  moisture 
is  soon  covered  with  a  fine  growth  of  a  whitish  or 
greenish  tint ;  this  is  formed  by  the  moulds,  lower  fungi 
belonging  to  the  vegetable  kingdom.  Under  the  microscope  a 
particle  of  this  growth,  teased  in  a  drop  of  water,  shows  a 
felt-work  of  tubes  or  filaments,  sometimes  segmented  at 
intervals,  sometimes  not ;  this  felt-work  is  the  mycelium.  From 
the  mycelium  little  stalks  stand  up  vertically  bearing  a  head 
which  itself  carries  little  granules  resembling  fruit.  In  the 
Mucors  the  head  is  a  spherical  sac  stuffed  full  of  these 
granules.  In  Penicillium  the  head  consists  of  ramifications 
or  digitations  dividing  in  their  turn,  the  last  sections  pinching 
off  little  pieces,  thzconidia;  the  whole  resembles  a  little  brush. 
In  Aspergillus  the  fructification  resembles  the  flower  of  an 
onion. 

A  fragment  of  mycelium  put  into  a  suitable  medium 
reproduces  the  mycelium.  The  conidia  can  do  the  same. 
In  the  case  of  the  mucorines  two  cells  can  join,  fuse,  and 
produce  an  "  egg " ;  often  these  two  cells  are  different,  and 
the  reproduction  may  be  regarded  as  sexual.  If  mucor  is 
grown  at  the  bottom  of  a  sugary  fluid  it  no  longer  forms  an 
abundant  mycelium ;  on  the  contrary  there  can  be  found 
nothing  but  short  pieces,  round  or  ovoid,  which  reproduce 
themselves  by  budding  like  yeasts.  But  any  of  these  pieces 
put  in  a  sugar-containing  medium  in  contact  with  the  air, 
forms  a  mycelium  on  the  surface.  This  transformation  of  a 


FORM  AND  STRUCTURE  OF  MICROBES     53 


FIG.  1 6. — Penicillium. 


FIG.  17. — Aspergillus. 


FIG.    1 8. — Mucor  taking   on   the  yeast  form 
When  grown  at  the  bottom  of  a  sugar  solution. 


FIG.  19. — Mucor. 


54 


MICROBES   AND   TOXINS 


mycelium  into  a  yeast-like  form,  which  also  produces  fermenta- 
tion like  a  true  yeast,  was  discovered  by  Pasteur,  who  thought 
that  in  the  course  of  ages  the  yeasts  had  been  evolved 
from  moulds  which  had  become  adapted  to  the  fermentative 
life. 


FIG.  20. — The  yeast  of  beer:  high 
or  tumbling  yeast. 


FIG.  21. — Spindle-shaped 
yeast. 


FIG.  22.— The  yeast  of 
wine. 


FIG.  23. — The  yeast  of  beer :  low 
or  heavy  yeast. 


Yeasts. — Yeasts  are  fungi  of  the  Blastomycete  group. 
They  consist  of  round  or  elliptical  cells  which  multiply  as  a 
rule  by  budding ;  some  multiply  by  equal  division  into  two 
parts.  Yeasts  may  furnish  spores,  which  number  2  to  10, 
and  may  remain  enclosed  in  the  mother  cell  until  by  the 
bursting  of  this  latter  their  germination  becomes  possible. 


FORM  AND  STRUCTURE  OF  MICROBES     55 

Bacteria. — The  term  "  microbes "  in  current  language 
most  frequently  signifies  bacteria.  The  bacteria  belong  to  the 
lower  plants,  are  unicellular,  contain  no  chlorophyll,  and  are 
almost  all  incapable  of  taking  up  carbon  from  the  carbonic 
acid  of  the  air.  They  are  incapable  of  life  except  in  the 
presence  of  ready  made  organic  matter,  and  are  in  consequence 
confined  either  to  a  saprophytic  existence  (moulds  on  a  fruit) 
or  to  a  parasitic  (as  in  the  case  of  the  typhoid  bacillus  in  the 
intestine).  Scattered  as  they  are  throughout  nature  the 
bacteria  are  the  chief  agents  in  the  decompositions  and  re- 
combinations of  organic  and  living  matter. 

The  usual  classification  of  bacteria  depends  on  their  external 
form  and  is  merely  provisional.  The  round  bacteria  are  called 
micrococci  or  cocci.  The 

streptococci  are,  as  Pas-       *                       *       f_      6 
teur  described  them,  like      * 
rosaries  or  necklaces ;  the 
staphylococci     are     like 
bunches  of  grapes.     Ac- 
cording as  the  micrococci  Sl*Q  .  -g        ^ 

in     their     multiplication 

divide   in   two    or    three       lg*  Q*^. 

planes  in  space,  they  ap-  cocci. — 4.   Staphylococci. — 5.    Sar- 

pear  arranged  in  the  mul-  S"1?/-^' 

o.   Vibrios.— 9.  opinlla. — 

berry  form  or  in  the  form  chaetes.— 11.  Clostridium. 

of  a  sardna  or  woolpack. 

The  bacilli  are  long  bacteria,  the  ends  of  which  are  sharply 

cut;  the  bacterium,  properly  speaking,  has  the  ends  rounded 

like  a  spindle  or  shuttle.     The  curved  bacteria  are  known  as 

vibrios,  spirilla,  and  spirochaetes. 

The  bacterial  cell  is  clothed  by  a  membrane  and  is  not 
naked  like  an  amoeba.  It  is  this  sheath,  resistant  and  elastic 
up  to  a  certain  point,  which  permits  a  corkscrew  spirillum  to 
maintain  its  form  in  spite  of  its  movements,  and  the  motile 
and  flexible  bacteria  to  recover  their  shape  after  distortion. 
The  membrane  takes  on  colours  different  from  the  protoplasm. 
It  is  not  of  the  same  composition  in  all  the  bacteria.  Some- 


56  MICROBES   AND  TOXINS 

times  it  is  of  cellulose,  sometimes  it  resembles  chitin,  some- 
times it  is  impregnated  with  fats  or  waxes  like  the  bacillus 
tuberculosis.  In  the  membrane  of  certain  bacteria  iron  and 

silicon  have  been 
found.  Sometimes 

in  °ld  cultures 

one  can  see  mem" 

branes  without 
typhoid  V&rto  Of  Tetanus       contents  which  re- 

Baxi&tts  QtoUlU  BatittttS      semble     empty 

FIG.  25.— Flagella  of  bacteria.  sheaths. 

Certain  bacteria 

possess  besides  Brownian  movement  an  independent  motility ; 
they  can  be  seen  performing  long  journeys,  pirouetting 
on  their  own  axis.  It  has  been  calculated  that  the  cholera 
vibrio  can  travel  18  centimetres  in  the  hour,  which  is 
equivalent  to  ten  or  fifteen  times  its  length  in  a  second. 
This  motility  is  due  to  cilia  or  flagella  analogous  to  those 
of  the  vibratile  epithelial  cells  or  the  ciliated  infusorians.  They 
are  often  longer  than  the  bacterium  which  carries  them.  In 
old  cultures  they  become  detached  and  scattered  through  the 
medium.  It  is  doubtful  whether  they  are  fine  expansions  of 
cellular  protoplasm  which  have  passed  through  pores  in  the 
enveloping  membrane  or  whether  they  are  expansions 
of  this  membrane  itself;  whether  they  are  not  mere 
propelling  organs  but  rather  tentacles  of  some  sort  which 
increase  the  surface  of  the  bacterium  and  thus  play  a  role  in 
nutrition,  especially  in  the  young  bacteria.  No  definite 
reply  can  be  given  to  these  questions.  According  to 
G.  de  Rossi's  experiments  on  agglutination  and  immunisation 
the  cilia  behave  like  protoplasm.  Their  action  ceases  at  very 
low  temperatures  and  above  50°  C.  When  a  bacterium 
provided  with  cilia  only  at  one  end  (certain  vibrios)  is 
attracted  towards  a  definite  substance  (positive  chemiotaxis), 
the  cilium  or  cilia  take  the  front  position  and  mark  the  head- 
end of  the  bacterium — just  as  the  flagellum  does  in  the  case  of 
the  trypanosomes. 


FORM  AND  STRUCTURE  OF  MICROBES     57 

Certain  bacteria  such  as  the  pneumococcus  and  the 
pneumobacillus  are  surrounded  by  a  sort  of  case  or  capsule. 
One  capsule  may  enclose  several  bacteria :  when  a  large  mass 
of  bacteria  is  included  in  one  enormous  capsule  what  is  called 
a  zooglcea  is  formed.  Bacteria  which  appear  encapsulated  in 
the  albuminous  fluids  of  the  infected  body  do  not  always 
possess  them  in  artificial  culture.  The  capsule  appears  to  be 
a  defensive  secretion.  The  anthrax  bacillus  defends  itself 
against  the  toxic  action  of  the  serum  of  the  rat  by  surrounding 
itself  with  a  thick  sheath  made  up  of  a  sort  of  mucus  which 
fixes  and  renders  harmless  the  toxin.  In  the  blood  of  lizards, 
which  are  very  resistant  to  anthrax,  the  bacillus  surrounds 
itself  with  a  thick  mucous  envelope.  The  streptococci 
occasionally  acquire  in  the  body  of  resistant  guinea-pigs  a 


FIG.  26. — Streptococci  surrounded  by  a  sheath  of  mucus. 

similar  defensive  covering.  The  streptococci  of  chronic 
infections  (otitis  and  nasal  infections)  are  frequently  encap- 
sulated. The  sarcinae  are  not  usually  pathogenic  but  encap- 
sulated sarcinae  have  been  described  which  were  pathogenic 
for  laboratory  animals. 

Bacteria  multiply  by  transverse  divisions,  Pasteuria  ramosa, 
described  by  Metchnikoff,  being  an  exception.  One  bacterium 
put  on  a  suitable  medium  produces  after  a  certain  time  two, 
these  two  producing  four,  and  so  on  ;  it  is  easy  to  imagine  to 
what  enormous  numbers  this  geometrical  progression  may  lead. 
In  nature  and  in  artificial  culture  media,  multiplication  is 


58  MICROBES   AND  TOXINS 

limited  by  the  quantity  of  available  nourishment  and  by  the 
accumulation  of  the  products  of  excretions  ;  the  cultures  end 
by  poisoning  themselves.  But  even  in  a  drop  of  culture 
medium,  the  energy  of  multiplications  is  enormous.  One 
filament  of  the  Bacillus  ramosus  studied  by  Marshall  Ward 
doubled  its  length  in  thirty-five  minutes ;  in  twelve  hours  a 
single  bacillus  produced  four  millions,  and  one  piece  of  one- 
hundredth  of  a  millimetre  in  length  produced  in  twelve  hours 
the  equivalent  of  a  thread  of  40  metres.  Pasteur  followed 
under  the  microscope  the  growth  of  the  yeast  of  wine  in  grape 
juice  at  13°  C.  One  globule  produced  10  millions  in  twenty- 
four  hours  when  nothing  intervened  to  limit  its  development. 
It  is  easy  to  understand  now  how  the  infinitely  minute  grows 
to  form  a  mass  and  brings  into  play  enormous  forces. 

Spores. — Certain  bacteria  produce  spores,  the  spore  appearing 
as  a  shining  point  in  the  filament  ;  the  protoplasm  of  the  cell 

diminishes  as  the 
spore  increases,  as 
if  speculations 
were  a  condensa- 
tion of  the  living 
matter.  Later  the 

rWUZ&tS  CftfoSpon  bacillus  disappears 
and  the  spore  is 
free.  Being  en- 
closed in  a  resist- 
ing sheath  the 
FIG.  27. — Various  types  of  germination  of  spores.  i  i 

I/The   spore   germinates   by  growth  hi  all        SPOre  resembles  a 
dimensions.     2.  Germination   by  a    sort   of       Seed,     capable    of 
terminal  budding.    3.  The  spore  germinating       Drolonged    D  r  e  - 
by  a  sort  of  lateral  budding.    (After  De  Bary 
and  Prazmowski.)  servation    and     of 

sprouting   into    a 

new  bacillus  when  the  conditions  become  favourable.  It  is 
by  means  of  their  spores  that  the .  bacilli  of  tetanus  and  of 
anthrax  persist  so  tenaciously  in  nature.  The  anthrax  bacillus 
is  killed  by  heating  to  60°  C.,  the  pore  not  till  after  three 
minutes'  boiling  at  100°  C, 


FORM  AND  STRUCTURE  OF  MICROBES     59 

The  spores  which  develop  in  the  interior  of  bacteria  are 
endospores.  The  name  of  arthrospores  has  been  given  to  those 
rounded  granules  which  certain  bacteria  produce  by  segmenta- 
tion, by  a  sort  of  pinching  off,  or  perhaps  by  a  shrinking  of 
their  substance  ;  for  example  the  cholera  vibrio,  which  does  not 
possess  endospores. 

The  properties  of  an  anthrax  bacillus  are  fixed,  and 
transmitted  to  the  new  generation,  by  a  sort  of  heredity,  by 
means  of  the  spore. 

It  is  not  impossible  that  some  sexual  differentiation  exists  in 
the  sporulating  bacteria. 

Pleomorphism  of  Bacteria. — The  bacterial  forms  have 
not  the  immutability  of  crystals,  nor  even  the  relative  stability 
of  species  among  the  higher  plants  and  animals.  They  are 
variable  to  an  extent  that  absolutely  confounds  the  bacteriolo- 
gist. In  a  pure  culture  such  a  diversity  of  forms  may  appear, 
forms  long  and  short,  rounded  and  thread-like,  that  the  culture 


FIG.  28. — Involution  forms  :  I.  Lactic  bacilli. — 2.    Bacillus  subtilis. — 
3    Anthrax  bacilli. 

might  be  thought  impure.  The  bacillus  of  diphtheria  trans- 
ferred from  a  natural  to  an  artificial  medium,  or  from  one 
artificial  medium  to  another,  appears  short  or  long  according 
to  circumstances  and  gives  in  old  cultures  forms  which 
resemble  staphylococci.  One  bacterium  exists  so  variable 
that  it  has  been  called  Proteus,  the  protean  bacillus.  The 
Bacillus  prodigiosuS)  the  bacterium  which  gives  red  colonies 


60  MICROBES   AND  TOXINS 

and  thus  produces  the  miracle  of  the  bleeding  host,  multiplies 
very  quickly  in  alkaline  media,  producing  coccal  forms  ;  these 
cocci,  reinoculated  on  a  faintly  acid  medium,  multiply  less 
rapidly  and  produce  bacillary  forms.  Bacteria  are  then 
multiform  or  pleomorphic  organisms. 

In  old  cultures  or  in  cultures  on  unfavourable  media,  forms 
are  seen  diverging  from  the  typical :  a  micrococcus  may  give 
relatively  enormous  globules  ;  a  bacillus  may  give  forms  racket- 
shaped,  club-shaped,  pear-shaped,  &c.  A  bacillus  which 
ordinarily  never  branches  may  show  branching  forms.  These 
abnormal  shapes  are  called  involution  forms,  again  examples  of 
pleomorphism. 

In  the  early  days  of  bacteriology  this  pleomorphism  was 
copiously  discussed.  Nageli  held  that  there  were  no  bacterial 
species,  but  that  the  bacteria  formed  an  immense  group  in 
which  neither  species  nor  families  nor  genera  were  to  be 
distinguished.  A  coccus  might  give,  under  suitable  conditions, 
a  rod  form  or  a  spirillum  and  the  function  might  be  as  variable 
as  the  form.  He  would  not  admit  that  a  given  fermentation 
always  corresponded  to  a  definite  bacterial  form,  but  held  that 
the  same  microbe  might  be  in  its  turn  an  acetic  ferment,  a 
lactic  ferment,  or  a  butyric  ferment.  For  him  bacterial  speci- 
ficity did  not  exist.  Hence  arose  the  belief  that  it  was  easy  to 
render  pathogenic  an  inoffensive  bacterium  like  the  B.  subtilis 
of  hay  infusions. 

The  botanist  Cohn  exaggerated  the  stability  of  bacterial 
species  as  much  as  Nageli  exaggerated  their  pleomorphism,  and 
bacteriologist-physicians,  in  their  desire  to  discover  for  each 
infectious  disease  a  specific  agent,  followed  Conn's  idea.  It 
was  obviously  impossible  to  hold  precise  opinions  about  the 
pathogenic  bacteria  if  there  existed  no  stability  in  their  forms. 
R.  Koch  accused  the  "  pleomorphists "  of  making  their 
observations  on  impure  cultures  or  on  infusions  containing 
already  a  very  numerous  flora ;  it  was  not  pleomorphism  that 
was  present,  but  confusion. 

Another  observer,  Kurth,  however,  demonstrated  the 
pleomorphism  of  the  Bacterium  Zopfii  in  pure  cultures ;  there- 


FORM  AND  STRUCTURE  OF  MICROBES     61 

upon  the  partisans  of  Cohn  and  Koch  declared  that  though  the 
saprophytic  species  might  lack  stability,  the  pathogenic  species 
were  stable.  In  their  eyes  the  bacillus  of  anthrax  was  always  a 
bacillus.  But  it  had  to  be  recognised  later  that  pleomorphism 
existed  even  among  pathogenic  species,  for  example  the  cholera 
vibrio  and  the  bacterium  of  fowl-cholera. 

Finally  the  stability  school  maintained  the  necessity  of 
distinguishing  between  the  constancy  of  the  mere  shape  and 
that  of  the  species — a  cholera  vibrio  may  modify  its  form  and 
yet  remain  always  the  vibrio  of  cholera — the  tadpole  of  the 
green  frog  does  not  resemble  an  adult  green  frog,  yet  it  belongs 
without  any  doubt  to  the  species  green  frog.  But  it  is  certain 
that  it  is  not  only  the  form,  but  also  the  species  qua 
species  which  is  variable  amongst  the  bacteria,  and  this 
variability  is  even  more  characteristic  of  their  physiological 
properties  than  of  their  shape.  The  bacillus  of  tuberculosis, 
amongst  others,  is  susceptible  of  very  divergent  adaptation. 

Bacterial  species  exist,  but  they  are  all  of  the  kind  called  in 
the  language  of  the  science  of  classification  "ill-defined" 
"That  the  methodical  classifier  should  be  frequently  embarrassed 
in  his  attempts  to  set  up  his  artificial  barriers,  is  very  far  indeed 
from  being  surprising.  The  world  was  not  created  for  the 
special  pleasure  of  descriptive  botanists,  and  it  is  equally  inter- 
esting to  science  to  possess  the  demonstration  that  a  classification 
is  impossible  as  to  have  established  a  classification,  had  such 
been  possible"  (E.  Duclaux). 

In  any  case  their  faculty  of  adaptation  and  their  power  of 
variation  sufficiently  explain  the  history  of  bacteria.  The 
pathogenic  species  must  have  come  from  saprophytic  species 
by  adapting  themselves  to  certain  animal  bodies.  The  same 
virus,  adapting  itself  in  the  course  of  ages  to  the  human  species 
and  to  the  cow,  has  produced  the  two  diseases  small-pox  and 
cow-pox,  and  this  adaptation  has  become  the  principle  of  a 
wonderful  prophylactic  procedure.  It  was  also  by  causing 
variations  in  the  virus  by  physical  and  chemical  action  that 
Pasteur  first  prepared  vaccines. 

More  recently  search  has  been  made  among  bacteria  for 


62  MICROBES   AND  TOXINS 

examples  of  mutation  resembling  those  studied  by  the  botanist 
Hugo  de  Vries.  In  a  pure  culture  of  bacillus  coli  Massini 
found  that  certain  individuals  suddenly  acquired  the  power  of 
fermenting  lactose,  and  that  this  property  was  transmitted  to 
their  descendants.  The  attenuation  of  the  anthrax  vaccines  of 
Pasteur  is  an  example  of  a  property  acquired  and  transmitted 
hereditarily  by  the  spore. 

Precisely  because  of  this  pleomorphism  and  plasticity  of 
bacteria,  one  must  take  care  not  to  imagine  mutations  too 
frequently.  Occasionally  modifications  may  be  got  which  are 
difficult  to  fix  and  which  do  not  constitute  true  races.  The 
bacillus  prodigiosus  has  been  cultivated  at  37*5°  C,  with- 
out producing  its  red  colour,  for  a  series  of  generations  ;  but 
after  the  thirty-fifth  passage,  when  put  again  at  22°  C.,  it  re- 
produced its  pigment.  Besides,  mutation  is  not  to  be  defined 
simply  as  a  sudden  variation.  There  must  be  transmission  by 
sexual  reproduction  in  addition.  One  could  not  speak  of 
mutation  except  in  cases  such  as  those  of  the  anthrax  bacillus, 
where  there  has  been  suspected  a  sort  of  sexuality  in  the  spore 
forms.  It  is  none  the  less  true  that  the  life  of  bacteria  presents 
numerous  facts  in  accordance  with  the  Darwinian  laws. 
"  Bacteriology,  like  all  branches  of  biology,  has  gained  by  the 
application  of  the  theory  of  evolution,  and  has  made  a  fair 
return  by  supplying  the  Darwinian  theory  with  a  striking 
confirmation  "  (Metchnikoff). 

The  Place  of  Bacteria  in  Classification.— 
Leeuwenhoek,  describing  in  1683  in  his  Arcana  naturae 
detecta  the  micro-organisms  of  the  mouth,  which  he  observed 
through  lenses  polished  by  himself,  represented  them  as 
animalcule.  In  1838  Ehrenberg  assigned  them  a  place  in  his 
work  on  the  Infusoria,  classing  them  with  the  Vibrios,  which 
he  regarded  as  animals.  But  their  evolution  and  their  activities 
indicate  that  the  bacteria  are  lower  plants. 

Are  they  to  be  classed  among  the  fungi  (moulds  and  yeasts) 
or  among  the  algae  ?  In  the  absence  of  chlorophyll,  bacteria 
resemble  fungi,  and  they  have  long  been  called  Schizomycetes, 
i.e.,  fungi  multiplying  by  transverse  division.  There  are 


FORM  AND  STRUCTURE  OF  MICROBES     63 


bacteria,  the  Streptothriceae,  which,  by  their  filamentous  and 
branching  appearance,  form  a  link  between  the  bacteria  and 
the  simplest  fungi.  On  the  other  hand,  there  are  yeasts  which 
resemble  bacteria  in  producing  endospores  and  multiplying  by 
division  instead  of  by  budding. 

It  is,  however,  with  certain  algae,  the  Cyanophyceae  or  blue 
algae  that  the  relationship  is  most  marked.  The  Cyanophyceae 
reproduce  by  division,  presenting  different  shapes,  long, 
round,  and  curved,  resembling  cocci,  coccobacilli  and  bacilli. 
They  are  pleo- 
morphic  like  the 
bacteria.  They 
possess  chloro- 
phyll in  contrast 
to  the  bacteria, 
but  the  pigment 
which  the  Cyano- 
phyceae possess 
in  addition  to 
their  chloro- 
phyll, the  phyco- 

chrome  (most  often  greenish-blue,  soluble  in  water  and 
diffused  throughout  the  cell)  is  not  without  some  analogy  to 
the  pigment  of  certain  bacteria.  Certain  algae  clump  together 
in  mucilaginous  sheaths  resembling  the  zoogloea  of  certain 
bacteria.  The  Cyanophyceae  do  not  form  endospores  but  have 
arthrospores  like  some  bacteria.  There  are  species  like 
Beggiatoa^  in  which  it  is  far  from  certain  whether  they  should 
be  classed  with  the  bacteria  or  with  the  Cyanophyceae.  These 
analogies  are  so  striking  that  a  family,  the  Bacteriacecz,  has  been 
made  for  the  algae  which  border  on  the  Cyanophycecz. 

But  in  view  of  the  differences  which  exist  between  these  two 
families  and  the  similarities  which  link  up  the  bacteria  and  the 
fungi  (moulds  and  yeasts),  certain  authorities  make  of  the 
bacteria  a  heteroclitous  group  where  there  are  brought  together 
representatives  more  or  less  altered  or  degenerated  of  various 
lower  plants.  There  exist  no  doubt  bacterio-algae ;  there  are 


FIG.  29. — Streptothrix.     Branching  bacteria. 


64  MICROBES    AND   TOXINS 

also   bacterial   moulds  and   bacterial   yeasts;   there  are  even 
protozoal  bacteria,  i.e.,  animal  bacteria,  the  Spirochsetes. 

The  conclusion  then  is  that  bacteria  are  not  the  original 
forms  from  which  the  others  were  derived  but  that  on  the 
contrary  the  bacteria  are  derived  from  the  more  definitely 
characterised  fungi  and  algae,  the  specific  characters  having 
become  obliterated  by  the  parasitic  habit.  The  branching 
forms  rarely  observed  in  the  bacillus  tuberculosis,  the  endo- 
spores  of  the  anthrax  bacillus,  the  arthrospores  of  the 
Streptothrices  are  all  atavistic  stigmata. 

The  Nucleus  of  Bacteria. — The  usual  nucleus,  consisting 
of  a  distinct  mass  in  the  body  of  the  protoplasm,  does  not  exist 
in  the  bacterial  cell. 

A  cell  without  a  nucleus  !  Several  observers  (A.  Fischer, 
Migula,  Massart)  do  not  shrink  from  such  a  paradoxical 
statement.  They  can  see  no  trace  of  a  nucleus  ;  what  others 
have  taken  for  it,  large  or  small,  is  only,  in  their  opinion,  an 
empty  space  or  a  vacuole,  taking  part  in  the  cell-division  and 
capable  of  dispersion  into  smaller  vacuoles.  The  scattered 
granulations  in  the  protoplasm  are  not  grains  of  chromatin, 
for  they  have  none  of  its  reactions.  They  are  either  products 
of  metabolism  or  reserve  materials.  They  have  been  called 
metachromatic  bodies,  because  when  stained  with  blue  or  violet 
stains  they  take  on  a  different,  reddish  tint. 

Such  observations  are  best  made  on  certain  large  bacteria 
found  in  nature.  It  is  remarkable  that  the  bacillus  asterosporus, 
which  was  the  one  selected  by  Migula  to  demonstrate  the 
absence  of  the  nucleus,  has  been  employed  by  others  to 
demonstrate  the  presence  of  one  or  even  several  well-defined 
nuclei.  Able  cytologists  complain  that  the  supporters  of  the 
nucleus  have  been  deceived  by  appearances  ;  that  they  have 
taken  for  chromatic  masses  the  transverse  segmentations  which 
appear  in  certain  large  bacteria  at  the  moment  of  division,  and 
that  they  have  even  chosen  for  their  demonstration  bacteria 
which  are  not  true  bacteria;  the  Bacterium  gammari  of 
Vejdowsky  is  said  to  be  a  fungus  approaching  the  yeasts  and 
multiplying  by  division,  while  another  bacterium  studied  by 


FORM  AND  STRUCTURE  OF  MICROBES     65 

the  same  author  in  the  digestive  tube  of  an  annelid  (Bryodrilus) 
ought  rather  to  be  considered  as  a  mould. 

To-day  one  can  agree  neither  with  those  who  affirm  that 
there  is  no  nucleus  nor  with  those  who  describe  a  well-defined 
nucleus.  In  reality,  the  bacteria  possess  a  diffuse  nucleus. 
Instead  of  forming  a  mass  with  sharp  contours,  the  chromatin 
is  scattered  through  the  protoplasm  in  the  form  of  granules 
quite  distinct  from  the  metachromatic  granules. 

Long  ago  Weigert  maintained  that  the  nucleus  is,  as  it  were, 
melted  or  dissolved  in  the  cytoplasm,  for  it  is  precisely  the 
nuclear  stains  which  stain  the  bacteria  best;  Biitschli  con- 
sidered that  the  cytoplasm  of  bacteria  is  reduced  to  a  thin 
layer  lying  in  apposition  with  the  membrane,  and  that  the 
bacterium,  far  from  lacking  a  nucleus,  is  in  reality  almost  all 
nucleus.  Further,  what  purpose  would  a  large  quantity  of 
cytoplasm  fulfil,  cytoplasm  whose  special  function  is  nutritive  ? 
Bacteria  are  parasitic,  and  absorb  their  nourishment  ready 
prepared.  Their  chief  business  is  to  multiply,  to  increase  in 
numbers ;  it  is  quite  natural  for  them,  therefore,  to  possess  an 
enormous  nucleus  like  the  spermatozoa  and  the  embryonic 
cells,  so  much  so  that  one  might  even  say  that  bacteria  are 
"  free  nuclei." 

Schaudinn  has  furnished  a  brilliant  support  to  these  ideas 
by  means  of  his  observations  on  the  largest  known  bacillus,  the 
Bacillus  Biitschlii)  found  by  him  in  the  intestine  of  cockroaches. 
The  chromatin  granules  are  scattered  through  the  protoplasm  ; 
to  produce  the  two  spores  they  come  together,  and  the 
chromatin  condenses  at  the  two  ends.  The  condition  of  a 
diffuse  nucleus  is  no  less  evident  in  the  B.  maximus  buccalis  of 
Swellengrebel. 

The  diffuse  nucleus  is  not  confined  to  bacteria.  There  are 
several  protozoa  which  normally  possess  a  well-defined  nucleus, 
but  at  certain  moments  in  development  or  in  certain  conditions 
of  nutrition  this  may  be  seen  changing  into  a  diffuse  nucleus. 
The  diffused  nuclei  have  been  called  by  R.  Hertwig  chromidia. 
Several  varieties  exist,  and  the  nucleus  of  B.  Butschlii,  like  the 
nuclei  of  bacteria,  in  general  should  be  considered  as  consisting 


66 


MICROBES  AND  TOXINS 


of  chromidia.  The  nuclei  of  the  Cyanophyceae  have  been  the 
subject  of  similar  disputes ;  in  these  inferior  algae  also  the 
existence  of  a  chromidial  nucleus  is  admitted,  quite  distinct 
from  the  metachromatic  bodies. 


p 

I 


•c 


ted 


FIG.  30. — Bacillus  maximus  buccalis.     (After  S wellengrebel. ) 

The  spiral  filament  represents  a  system  of  chromidia  in  process  of  division 

(4  and  5). 

Reproduction  and  Sex. — A  species  is  said  to  possess 
sex  when  fertilisation  occurs  by  the  fusion  of  two  nuclei 
differentiated  into  male  and  female. 

In  the  haematozoa  of  malaria  the  sexual  act  takes  place 
in  the  body  of  the  mosquito  by  the  conjunction  of  the 
flagellum  and  the  female  cell,  and  at  this  point  the  sexual  cycle 
begins. 

Since  Schaudinn's  discovery  of  the  trypanosome  forms  in 
the  cycle  of  certain  haematozoa  closely  related  to  those  of 
malaria,  one  must  admit  the  existence  of  male  and  female 
forms  in  the  case  of  trypanosomes.  Besides  the  asexual 
multiplication  by  longitudinal  division  there  would  thus  exist 
among  trypanosomes  true  sexual  reproduction. 

Among  the  fungi  the  ovum,  the  complete  reproductive  form, 


FORM  AND  STRUCTURE  OF  MICROBES     67 


results  from  the  fusion  of  protoplasm  as  well  as  nucleus  of 
two  differentiated  cells,  the  gametes,  one  male,  the  other 
female.  Heterogamy  exists  when  the  male  gamete  is  manifestly 


FIG.  31. — Successive  stages  of 
conjugation  in  a  Zygosac- 
charomycete.  (After  Barker.) 


FIG.  32. — Zygoscucharomyces  yeast. 
(a)  Vegetative  cells  :  (6)  Asci. 


different  from  the  female — isogamy  when  they  seem  identical. 
The  ascus  and  the  basidium  are  modes  of  sexual  reproduction. 
In  the  yeasts,  examples  of  conjugation  are  not  lacking. 
In  Zygosaccharomyces  before  the  intracellular  formation  of  the 
spores,  two  neighbouring  cells  put  out  each  a  little  bud ;  the 
two  buds  join  and  along  the  little  canal  thus  formed  fusion  of 
the  nuclei  takes  place  (according  to  Barker).  The  two  cells 


FIG.  33. — Nuclear   fusions  and   spore   formation    in   Schizosaccharomyces 
octosporus.     (After  Guilliermond. ) 

remain  united  forming  a  dumb-bell-shaped  ascus.  The  same 
process  occurs  in  certain  schizosaccharomycetes,  where  the 
shape  of  the  ascus  also  recalls  its  origin  from  two  cells.  Since 

F  2 


68  MICROBES    AND   TOXINS 

the  cells  which  join  belong  to  the  same  strain,  and  to  the  same 
generation  or  to  two  generations  very  near  each  other,  this 
conjugation  in  the  yeasts  is  an  example  of  isogamy. 

In  the  bacteria  it  has  long  been  observed  that  multiplication 
by  transverse  division,  /.<?.,  asexual  reproduction,  appeared  to 
be  an  absolute  rule;  but  Forster  has  described  a  species  of 
conjugation  among  the  sulphur-forming  bacteria;  they  come 
together  in  twos  or  threes,  put  out  little  processes,  unite  and 
finally  separate;  this  is  a  conjugation  between  individuals 
which  are  doubtless  differentiated,  and  resembles  somewhat 
the  well-known  conjugation  of  infusoria. 

The  observations  of  Schaudinn  on  B.  Biitschlii  have  demon- 
strated the  existence  of  conjugation  also  among  the  bacteria — 
at  least  among  those  which  produce  endospores.  The  ordinary 
multiplication  by  transverse  division  exists  in  B.  Butschlii,  but 
in  the  individual  about  to  form  spores,  other  peculiar 
phenomena  are  seen  :  the  individual  begins  by  dividing  into 
two,  but  the  septum  disappears  almost  as  soon  as  it  is  formed 
and  is  dissolved,  the  two  cells  which  had  just  been  divided 
by  the  septum  melting  again  into  a  single  cell ;  an  exchange 
of  chromatin  takes  place  between  them,  and  finally  almost  all 
the  chromatic  granules  collect  at  the  two  poles  to  form  two 
spores.  Short  as  is  the  time  that  the  septum  remains,  there 
have,  nevertheless,  existed,  during  that  moment,  two  cells 
which  have  conjugated :  and  this  is,  according  to  Schaudinn, 
a  rudimentary  sexual  act.  An  analogous  conjugation  occurs 
with  B.  Sporenema. 

It  is  really  an  autogamy  or  fusion  between  two  elements  of 
the  same  cell,  or  to  be  more  precise  it  is  a  conjugation  between 
two  daughter-cells  of  the  same  mother-cell,  and  is  hence  called 
paedogamy. 

Numerous  cases  of  autogamy  have  been  described  among 
the  protozoa.  Autogamy  is  the  opposite  of  the  amphigamy 
which  occurs  when  there  is  conjugation  of  two  individals  of 
well-differentiated  sex. 

What  relations  are  to  be  established  between  these  two 
methods  of  fertilisation?  Is  autogamy  the  simple  primitive 


FORM  AND  STRUCTURE  OF  MICROBES     69 

type   from   which   the   other   is   derived,    or   is    it   simply  a 
degenerated  degraded  form  ?    As  far  as  bacteria  are  concerned, 


FIG.  34. — Bacillus  Biitschlii.  (After  Schaudinn.)  I  to  6.  stages  of  trans- 
verse division  without  sporulation.  —7  to  9.  Rudimentary  sexual 
process. — 10  to  14.  Spore  formation.  — 15  to  16.  Spore  germination. 

Schaudinn  regarded  the  conjugation  which  exists  in  B. 
Biitschlii  as  an  evidence  of  sex,  though  rudimentary,  degraded 
and  residual.  Without  absolutely  excluding  the  possibility  of 


70  MICROBES   AND   TOXINS 

primitive  autogamy  the  Schaudinn  school  considers  autogamy 
in  general  as  a  regressive  type. 

The  truth  is  that  sexual  differentiation  of  the  gametes  is 
universal  in  nature  even  in  the  lower  orders.  Researches  on 
the  Protista  permit  the  recognition  of  sexual  dimorphism  in 
every  nucleus,  i.e.,  there  is  in  every  nucleus  a  double  substance 
and  a  double  function,  one  specially  nutritive,  the  female,  the 
other  specially  reproductive,  the  male.  Every  protozoon  cell 
is,  to  some  extent,  hermaphrodite,  but  with  one  or  other 
element  predominating.  Even  in  autogamy,  the  elements 
which  fuse  are  differentiated. 

Sex  is  thus  universal  in  nature,  and  it  is  from  this  point  of 
view  that  Schaudinn  had  to  consider  the  rudimentary  con- 
jugation of  B.  Butschlii  as  a  degenerate  type.  As  with  the 
structure  of  the  nucleus,  the  fact  of  autogamy  indicates  again 
that  bacteria  belong  to  an  order  degraded  by  the  parasitic 
habit. 

Chemical  Composition. — The  chemical  composition 
varies  with  the  species  studied,  and  in  the  same  species  with 
the  age  and  the  nutriment.  Duclaux  observed  in  the  cells  of 
a  fifteen-year-old  yeast  that  the  proportion  of  fatty  material, 
in'stead  of  being  5  per  cent,  as  in  young  yeasts,  had  risen  to 
20-30  or  even  52  per  cent,  the  proportion  of  nitrogen  varying 
from  i  to  4. 

Bacterial  substances  contain  proteids,  fats,  and  sugars,  but 
as  there  are  so  many  proteids  and  so  many  sugars,  the  figures 
can  only  give  a  general  indication.  The  cell  is  a  "  well  stocked 
laboratory,"  and  one  which  is  always  in  activity. 

The  organic  materials  isolated  from  bacterial  bodies  of  very 
diverse  species  are  the  following  : — 

Coagulable  albumins  in  the  expressed  juices  ;  globulins  ;  a  protamine  (from 
B.  tuberculosis)  ;  proteoses  (by  peptic  digestion). 

Glycoproteids.  Phosphoproteids  and  their  derivatives  :  nucleins,  nucleic 
acids,  xanthin  bases,  and  pyridin  bases. 

A  substance  resembling  chitin  or  keratin  (B.  tuberculosis). 

Products  of  hydrolysis  of  proteins  :  amino-acids  and  hexone  bases. 

Carbohydrates  :  Cellulose,  hemicellulose,  and  sugars. 

Fats  and  waxes,  neutral  fats  (capsulated  bacteria,  B.  diphtheria,  B.  tuber- 
culosis). Free  fatty  acids  (B.  tuberculosis)  ;  waxee  (B.  tub.)  ;  lecithin 


FORM  AND  STRUCTURE  OF  MICROBES     71 

(B.  tub.,  1 6  per  cent.,  according  to  Kressling) ;  phosphorized  fats  ;  in 
cultures  of  B.  tuberculosis  of  a  certain  age  the  content  of  fat  and  wax 
varies  between  20  and  39  per  cent. 
Ash  :  B.  tuberculosis  8  per  cent.,  B.  coli  8*5  per  cent. 

Ruppel  gives  the  composition  of  the  Bacillus  tuberculosis  as 
follows : 

Tuberculinic  acid      8*5  per  cent. 

Nucleoprotamine       «     .«     24*5       „ 

Nucleoproteid    «     23          „ 

Fats  and  waxes ...     ...      26*5       „ 

Ash      M     «     ...     9*2      „ 

Proteids  (keratin)      «     8-3      „ 

Most  of  these  substances  pass  into  the  extract,  which  is 
known  as  tuberculin. 

For  comparison  :  the  human  body,  taken  as  a  whole,  con- 
tains 65  to  70  per  cent,  of  water,  the  plants  used  for  food  60  to 
80  per  cent,  algae  90  per  cent. 

cteria  are  specially  rich  in  albuminoids.  The  moulds 
contain  more  carbohydrate  than  nitrogenous  material,  their 
outer  covering  containing  cellulose. 

Yeasts  contain  much  nuclein ;  as  they  grow  older  the  fat 
content  increases  ;  their  outer  covering  also  is  cellulose.  The 
mineral  matter  consists  of  phosphoric  acid,  potassium,  mag- 
nesium, sodium,  silicon,  lime,  sulphur,  and  oxide  of  iron. 
Yeast  cells  are  richer  in  glycogen  than  the  liver  cells  of  the 
rabbit  (31  and  10  per  cent,  of  the  weight  of  dried  yeast). 


CHAPTER   IV 

PHYSIOLOGY   OF   THE    MICROBES 

Nutrition— The  definition  of  food  material — Nutrition  of  the  Mucedinese — 
Raulin's  experiments — Bearing  of  these  experiments — Nutrition  of 
yeasts  and  bacteria — Importance  of  the  chemical  constitution  of  the 
medium — The  bacterium  of  sorbose — The  idea  of  the  'soil' — Respira- 
tion— Aerobic  and  anaerobic  life. 

The  purple  bacteria. 

Secretion  of  diastases  or  enzymes. 

Products  of  the  cultures — Products  of  excretion. 

Auto-intoxication  of  the  cultures. 

Production  of  heat — Production  of  light — Production  of  pigment. 

Action  of  heat  on  microbes  ;  thermophil  microbes. 

Action  of  light ;  ultra-violet  rays. 

Physiology  of  Protozoa :  cells  possessing  a  great  abundance  of  diverse 
functions — much  differentiated — Nutrition  ;  digestion,  respiration 
Irritability — Reproduction — Parasitism — Adaptations  and  specificities 
— Life  cycle  and  secondary  host — Cultures. 

THE  microbes  act  in  nature  as  transformers  of  energy.  The 
energy  which  they  get  from  oxygen  and  from  various  food 
materials  they  transform  into  products  of  excretion,  heat, 
light,  and  work,  retaining  a  portion  used  up  in  the  building  of 
their  own  substance.  Like  the  higher  creatures  they  display 
anabolism  and  catabolism.  Each  one  is  like  a  little  vortex, 
from  which  the  organic  matter  emerges  different  from  its  state 
on  entry.  It  is  characteristic  of  living  beings  to  transform  the 
molecules  of  their  food  materials  into  more  complicated 
molecules.  For  example,  from  sugar  they  produce  cellulose, 
from  carbohydrates  along  with  the  nitrogen  of  ammonia  they 
prepare  the  nitrogenous  compounds. 

To  construct  these  new  arrangements  of  the  molecule,  energy 

72 


PHYSIOLOGY   OF  THE    MICROBES          73 

or  heat  is  necessary,  and  the  microbe  provides  this  by  burning 
up  a  portion  of  its  food  material.  Thus,  while  part  of  the  food 
is  raised  to  a  level  of  higher  organisation,  another  portion,  on 
the  other  hand,  is  reduced  to  a  simpler  condition.  For 
example,  part  becomes  protein,  part  carbonic  acid  and  water, 
the  latter  not  being  incorporated  in  the  living  substance. 

Similarly,  a  microbe,  like  the  Aspergillus  niger,  burns  up 
with  the  help  of  the  oxygen  of  the  air  the  sugar  which  is 
furnished  by  the  food,  and  this  combustion  puts  at  its  disposal 
a  certain  number  of  calories  ;  one  molecule  of  sugar  weighing 
1 60  grams  furnishes  673  calories  during  its  transformation  into 
CO.,  and  H2O.  The  yeast  of  beer,  which  decomposes  sugar 
less  completely,  *.*.,  into  alcohol  and  carbonic  acid,  only  yields 
33  calories.  If  the  yeast  had  the  nutritive  requirements  of  the 
aspergillus  it  would  have  to  use  up  nearly  twenty  times  more 
food.  Nutrition  of  this  kind  is  characteristic  of  a  ferment, 
and  the  fermentative  power  is  greater  the  more  of  the  food- 
stuff the  cell  is  obliged  to  break  down. 

Lacking  chlorophyll  as  they  do  almost  universally,  the 
microbes  cannot  take  up  carbon  directly  from  the  air  as  do 
the  green  plants.  They  demand  their  food  ready  made  so 
that  they  can  destroy  it,  turning  it  into  carbonic  acid  and 
water.  The  heat  derived  from  this  destruction  takes  for  them 
the  place  of  the  energy  furnished  by  sunlight  to  the  plants 
which  contain  chlorophyll. 

The  food  of  a  microbe  may  then  be  denned  as  "every 
material  from  which  a  given  microbe,  under  the  conditions  of 
the  experiment,  can  take  the  material  necessary  for  its  develop- 
ment and  the  heat  necessary  to  render  it  independent  of  solar 
energy."  In  calculating  the  total  energy  coming  into  play 
in  the  protoplasmic  activity  a  certain  amount  has  to  be 
accounted  for  as  external  heat,  and  it  is  even  the  rule  for  an 
excess  to  appear  as  heat,  so  that  there  is  a  rise  of  temperature 
in  the  medium.  Thus  a  vat  at  the  surface  of  which  acetic 
fermentation  is  going  on  gets  notably  hotter,  and  the  same  is 
the  case  in  grapes  undergoing  vinous  fermentation.  But 
under  certain  limited  conditions  the  protoplasm  can  perfectly 


74  MICROBES   AND  TOXINS 

well  employ,  as  part  of  its  food,  substances  already  oxidised 
and  incapable  in  any  way  of  furnishing  heat,  the  conditions 
being  that  these  are  made  to  enter  into  a  nutritive  compound 
in  the  manufacture  of  which  heat-furnishing  transformations 
occur  in  sufficient  amount.  For  example,  the  nitric  bacteria, 
as  Winogradsky  has  shown,  can  take  their  carbon  from 
carbonic  acid  on  condition  that  at  the  same  time  they  trans- 
form nitrous  acid  into  nitric.  Similarly,  the  ferments  which 
fix  nitrogen  can  take  up  this  gas  from  the  air  on  condition 
that  they  destroy  at  the  same  time  by  oxidation  sugar  or  some 
other  hydrocarbon  capable  of  furnishing  heat  during  oxidation." 
(E.  Duclaux). 

Cellular  protoplasm  elaborates  food  by  means  of  the 
diastases  which  it  contains,  and  has  peculiar  wants  and 
preferences  according  to  the  species.  It  must  therefore  be 
difficult  for  a  microbe  to  find  the  particular  nourishment  which 
suits  it  best.  The  artificial  cultures  of  microbes  which  are 
so  useful  in  scientific  research  and  in  medicine  demand 
practically  for  each  microbe  those  food-stuffs  which  nourish  it 
in  nature. 

Nutrition  of  the  Mucedinese  (Raulin's  Experi- 
ments).— Raulin  succeeded  in  composing  with  perfectly  pure 
sugar  and  mineral  salts  an  artificial  medium  more  favourable 
to  the  growth  of  Aspergillus  niger  than  any  occurring  in 
nature.  It  is  thus  a  cultivation  in  the  fullest  sense  of  the 

word. 

Raulirfs  Medium. 

Water ideograms. 

Candy-sugar 70  ,, 

Tartaric  acid      4  ,, 

Ammonium  nitrate „     4  ,, 

,,         phosphate 0*60  gram. 

Potassium  carbonate         „     ...     ...  O'6o  ,, 

Magnesium      „               «     ....  0*40  ,, 

Ammonium  sulphate        ...     ... «     0*25  ,, 

Zinc                    „              0-07  ,, 

Ferrous              „ ....     ...  0*07  ,, 

Potassium  silicate             ...     ....     .„     0*07  „ 

This  medium  was  prepared  by  a  series  of  trials  and 
demanded  a  most  admirable  patience,  for  he  had  to  make 


PHYSIOLOGY   OF  THE   MICROBES         75 

modifications  in  the  number  and  quantity  of  the  constituents, 
to  compare  the  weights  of  the  growth  obtained,  to  determine 
the  temperature  and  the  hygrometric  condition  of  the  atmos- 
phere, and  even  to  take  account  of  the  shape  of  the  culture 
flasks,  which  affected  the  oxygen  supply. 

If  potassium  is  cut  out  from  the  above  formula,  the  crop  of 
aspergillus  is  25  times  less,  all  the  other  conditions  remaining 
the  same;  if  the  figure  25  is  taken  as  representing  the 
measure  of  utility  of  potassium,  the  utility  of  the  other  food- 
stuff may  be  represented  as  follows  :  nitrogen,  153  ;  phosphorus, 
182;  sulphur,  25;  silicon,  1*4;  magnesium,  91;  zinc,  10; 
iron,  27. 

If  you  take  the  ratio  between  the  weight  of  one  of  these 
food-constituents  and  the  weight  of  the  crop  due  to  its 
presence,  you  get  a  number  which  expresses  its  specific  utility, 

e.g.,  for  zinc  ^  =  560. 

This  ratio  was  in  several  experiments  found  to  be  953. 
For  nitrogen  it  is  17;  for  phosphorus  157;  sulphur,  346; 
potassium,  64;  magnesium,  200;  iron,  857.  It  is  to  be  noted 
that  aspergillus  takes  up  zinc  from  a  medium  in  which  the  zinc 
is  diluted  i  in  50,000. 

If  i  in  1,600,000  of  silver  nitrate  is  added  to  the  fluid, 
aspergillus  spores  no  longer  sprout  in  it :  even  when  simply 
poured  into  a  silver  vessel  the  fluid  dissolves  sufficient  silver 
to  prevent  growth.  The  plant  is  thus  an  indicator  so  sensitive 
as  to  mark,  by  its  refusal  to  sprout,  such  small  quantities  as 
i/24oth  of  sulphate  of  copper,  i/8,oooth  of  platinum  bichloride, 
i/5o,oooth  of  perchloride  of  mercury. 

Zinc  has  a  definite  food  value  but  iron  acts  differently, 
namely,  by  neutralizing  a  substance  which  is  produced  by  the 
growth  and  becomes  injurious  to  it,  perhaps  sulphocyanic  acid. 

Tartaric  acid  maintains  the  acidity  of  the  medium  and 
prevents  bacteria  from  contaminating  the  culture,  for  almost 
all  bacteria  refuse  to  grow  except  in  an  alkaline  medium. 
Hence  a  culture  of  aspergillus  in  Raulin's  fluid  produces  its 
own  asepsis.  The  sugar  is  only  assimilated  after  inversion  by 


76  MICROBES   AND   TOXINS 

a  diastase  of  the  fungus  ;  three  parts  by  weight  of  sugar  furnish 
about  one  of  growth.  When  glucose  is  supplied  instead  of 
saccharose,  the  culture  starts  off  more  quickly ;  lactose  is  not  a 
good  food-stuff.  Alcohol  (in  quantity  equivalent  to  the  weight 
of  sugar)  interferes  with  the  germination  of  the  spores,  but  in 
the  adult  form  the  fungus  gets  on  quite  well  with  it ;  alcohol 
is  thus  a  poison  to  the  embryo,  but  good  food  for  the  adult. 
Starch  paste  (boiled)  can  supply  the  necessary  carbohydrate 
nourishment,  but  raw  starch  alone  prevents  germination ;  the 
adult  would,  however,  attack  it  as  a  sort  of  last  resource. 

The  Bearing  of  Raulin's  Experiments. — These  are 
not  mere  laboratory  fantasies  but  are  in  reality  almost  the  first 
exact  experiments  on  the  conditions  of  plant  cultivation  and 
growth.  For  every  plant  in  the  fields,  for  every  microbe  in  the 
laboratory,  a  Raulin's  fluid  is  demanded,  a  fluid  which  would 
represent  ideal  conditions  for  development  :  all  the  improve- 
ments in  the  technique  of  cultivation  are  attempts  in  this 
direction.  For  example,  the  first  cultures  of  the  tubercle 
bacillus  were  made  with  great  difficulty  on  coagulated  blood 
serum  :  good  growth  was  only  obtained  by  adding  glycerine  to 
the  nutritive  medium  (Nocard  and  Roux).  Almost  all  our 
culture  media  for  bacteria  are  empirical :  blood,  ascitic  fluid, 
and  serum  are  supplied  to  them  because  we  know  that  they 
live  well  in  the  animal  body  in  these  fluids.  If  our  knowledge 
of  bacteria  was  as  far  advanced  as  is  that  of  Aspergillus  it 
would  be  possible  to  make  media  of  known  and  constant 
composition  with  measured  quantities  of  the  constituents,  and 
these  would  undoubtedly  be  of  the  greatest  value  in  the 
preparation  of  vaccines  and  toxins.  When  we  consider  that 
Aspergillus  is  sensitive  to  zinc  in  the  dilution  of  i  in  50,000 
and  to  silver  nitrate  in  the  proportion  of  i  in  1,600,000,  it  is 
possible  to  foresee  to  some  extent  the  solution  of  many 
technical  difficulties  and  to  appreciate  the  extent  of  what 
remains  to  be  discovered  in  connection  with  the  action  of 
manures,  food-stuffs  and  drugs.  Raulin's  experiments  show  to 
what  degree  bacteriology  and  medicine  depend  on  the  progress 
of  chemistry. 


PHYSIOLOGY   OF  THE   MICROBES         77 

We  know  how  to  prepare  a  clean  vaccine  fluid  by  culture 
on  the  flanks  of  the  calf,  but  we  cannot  prepare  in  vitro  a  vaccine 
lymph  bacteriologically  pure.  This  would  be  possible  if  we 
knew  the  nutritive  demands  of  the  vaccine  virus.  This  virus 
is  a  microbe  still  unknown,  but  probably  living  in  the  interior  of 
the  epidermal  cells  ;  it  finds  itself  there  in  a  medium  in  which 
reducing  actions  predominate.  Repin  succeeded  in  preparing 
a  reducing  medium  by  means  of  a  living  reducing  agent  put 
into  a  culture  flask  (the  tyrosinase  extracted  from  the 
mushrooms  of  the  genus  Russula) ;  with  this  he  got  a 
commencement  of  growth  in  the  vaccine  under  artificial 
conditions.  It  is  obvious  what  problems  lie  in  wait  for  those 
who  try  to  grow  bacteria  in  the  laboratory. 

The  Nutrition  of  Yeasts. — The  yeasts  are  of  such  great 
industrial  value  that  every  detail  of  their  nutrition  has  had  to 
be  studied.  They  demand  phosphoric  acid  and  potassium, 
magnesium,  lime,  and  sulphur;  their  nitrogenous  food  they  take 
from  ammoniacal  salts  and  they  can  use  those  albuminoid 
substances  which  are  soluble  in  water,  dialysable,  and  more  or 
less  insoluble  in  alcohol,  and  which  exist  in  serum  ;  also  they 
can  use  urea  and  allantoin.  The  carbohydrates  they  employ 
are  in  the  first  place  sugars,  then  various  alcohols,  acids  and 
organic  salts.  From  its  food  the  yeast  accumulates  reserve 
material,  i.e.,  glycogen.  Yeast  attacks  the  food-stuffs  supplied 
to  it  by  means  of  its  diastases  ;  and  in  doing  so,  while  toiling 
for  its  own  purposes,  it  toils  for  ours — exactly  like  a  hive  of  bees. 

Alimentation  of  Bacteria. — The  minerals  employed  in 
nutrition  are  rather  varied ;  sulphur,  phosphorus,  calcium, 
magnesium,  potassium,  sodium,  traces  of  iron,  traces  of  chlorine. 
As  carbohydrate  food,  sugars  and  glycerine ;  as  nitrogenous, 
ammoniacal  salts  and  peptones,  natural  proteins  like  blood 
serum  and  asparagine.  The  food-stuffs  are  supplied  by  meat 
infusions,  bouillons  with  peptone  and  salt,  by  animal  fluids  such 
as  serum,  urine,  ascitic  fluid,  milk,  and  by  fruit  juices.  The 
preferences  which  bacteria  show  for  certain  foods  are  employed 
for  diagnosis,  because  bacteria  are  characterised  not  less  by 
their  food  preferences  than  by  their  shape. 


78  MICROBES   AND   TOXINS 

Sulphur  is  indispensable  in  the  culture  fluid  of  the  sulphurous 
o.r  sulpho-bacteria  (Beggiatoa,  Lamprocystis,  and  other  species 
described  by  Winogradsky) ;  these  can  do  almost  entirely 
without  organic  food  material  and  grow  well  in  water  which 
contains  only  4*8  milligrams  of  this  per  litre,  but  which  contains 
two  milligrams  of  sulphuretted  hydrogen.  This  they  decompose, 
fixing  the  sulphur  and  accumulating  it  in  the  cell  in  the  same 
way  as  yeasts  with  glycogen.  Certain  Beggiatoa  contain 
80-95  Per  I0°  °f tnen"  weight  of  sulphur.  When  put  for  two  or 
three  days  in  non-sulphurous  water  they  oxidize  their  sulphur 
turning  it  into  sulphates.  If  the  dearth  of  sulphur  continues 
they  die. 

The  ferro-bacteria  (e.g.^  Crenothrix  polyspora^  Cladothrix 
dichotoma,  Leptothrix  ochraced)  oxidize  the  carbonate  of 
iron  protoxide,  FeH2(CO3)2,  and  accumulate  the  hydrate  of 
ron  oxide.  Instead  of  iron  oxide  a  deposit  of  oxide  of 
manganese  has  been  observed  in  certain  cases. 

The  Importance  of  the  Chemical  Constitution  01 
the  Medium. — Pasteur  observed  the  relations  between  the 
chemical  structure  of  the  food  material  and  the  physiological 
action  on  it  of  the  microbe.  A  Penicillium  uses  up  dextro- 
tartaric  acid,  leaving  intact  the  laevo  form  until  the  former  is 
completely  exhausted.  Penicillium  glaucum  and  certain 
yeasts  can  decompose  optically-inactive  sugars,  burning  up  the 
dextrorotatory  form  while  sparing  the  laevo. 

Following  in  the  track  of  Emil  Fischer,  there  have  been 
observed  relations  between  the  molecular  constitution  of  a 
sugar  and  its  value  as  food  or  fermentable  material  to  a  yeast 
or  in  general  to  any  definite  ferment.  Among  the  numerous 
sugars  with  the  general  formula  C^H^On  (n  being  a  whole 
number  i,  2,  3,  4,  etc.),  the  ordinary  yeasts  only  ferment  those 
with  the  carbon  atoms  numbering  3  or  a  multiple  of  3.  Sugars 
which  are  isomeric,  but  whose  molecule  does  not  possess  the 
same  stereochemical  configuration,  do  not  behave  exactly  the 
same  towards  a  given  yeast.  In  a  mixture  of  glucose  and 
levulose  one  or  other  is  the  first  to  be  decomposed,  this  varying 
with  the  strain  of  yeast. 


PHYSIOLOGY   OF  THE   MICROBES         79 

There  exist  also,  it  is  true,  "fermentations  by  force  of 
example,"  where  a  fermentation  started  on  a  certain  sugar  may 
extend  its  attack  to  another  sugar  which  at  first  was  not 
fermentable :  galactose,  for  example,  can  be  fermented  when 
it  has  been  "  baited "  by  glucose.  Similar  affinities,  capable 
of  modification  by  custom,  come  into  play  without  doubt  in  the 
sensitiveness  or  resistance,  natural  and  acquired,  of  an  animal 
body  towards  a  pathogenic  ferment.  Beneath  the  biological 
specificity  there  lies  a  chemical  specificity. 

One  of  the  best  examples  that  can  be  quoted  is  that  of  the 
bacterium  of  sorbose  studied  by  G.  Bertrand. 

This  makes  a  selection  among  the  polyatomic  alcohols, 
attacking  only  in  their  molecule  a  link  of  the  formula  CH'OH, 
transforming  it  into  CO  and  consequently  producing  always  a 
ketone  body.  Further,  for  this  link  to  be  attacked  its  hydroxyl 
OH  must  not  be  on  the  side  of  the  H  atom  of  the  neighbour- 
ing link  CHOH.  Finally  the  secondary  group  attacked  is 
always  next  to  one  of  the  primary  group  CH2OH,  which 
terminates  the  chain,  at  least  in  the  formulae  below  C7.  The 
stereochemical  structure  of  the  sugars  thus  plays  an  important 
part  in  the  matter  :  it  is  on  it  and  on  it  alone  that  the  possibility 
of  attack  by  the  bacterium  depends.  Its  action  is  so  narrowly 
specific  that  it  only  transforms  certain  chemical  groups,  taking 
no  interest  in  the  rest  of  the  molecule,  whatever  may  be  its 
mass  and  structure.  A  fermentation  of  this  kind  has  all  the 
value  of  a  chemical  reaction. 

Food-stuffs,  culture  media,  infected  organisms,  represent 
for  the  bacteria  their  soil,  "  If  we  consider,"  says  Bertrand, 
"on  the  one  hand  the  differences  in  chemical  composition, 
even  qualitative,  which  may  exist  between  two  closely  related 
species,  and  on  the  other  hand  the  extraordinary  variety  of 
proteins  which  it  is  possible  to  conceive  of  nowadays,  it  will 
hardly  appear  unreasonable  to  compare  animal  species,  or 
physiological  variations  of  the  same  species,  to  culture 
media  varied  like  those  which  I  used  in  the  study  of  the 
sorbose  bacterium,  nor  to  account  for  their  immunity  or 
susceptibility  towards  a  given  microbe  by  a  chemical  or  even 


80  MICROBES   AND   TOXINS 

merely  stereochemical  difference  in  their  composition."  In 
two  culture  media,  identical  except  that  one  contains  sorbite, 
the  other  dulcite,  a  different  sugar,  the  former  supports  the 
sorbose  bacterium,  the  latter  is  refractory.  To  the  bacillus 
tuberculosis  distributed  everywhere,  all  the  "  soils  "  are  not  the 
same.  The  chemical  study  of  the  soil  ought  to  go  hand  in 
hand  with  the  study  of  the  microbe. 

Respiration  :  Anaerobic  Life — Oxygen  is  the  primary 
food  of  creatures  which  have  respiration.  Lavoisier  showed 
that  oxygen  is  indispensable  to  life. 

During  his  study  of  the  fermentative  change  of  calcium 
lactate  into  butyrate,  Pasteur  discovered  the  vibrio  butyricus, 
and  made  the  fundamental  observation  that  this  organism  lives 
without  free  oxygen  and  even  dies  on  contact  with  the  air  : 

"  Pure  carbonic  acid  passed  for  however  long  a  period  through 
the  fluid  in  which  they  are  growing  has  no  effect  whatever  on 
their  life  and  multiplication.  If  atmospheric  air  is  passed 
through  instead  for  one  or  two  hours  under  precisely  parallel 
conditions  they  all  die,  and  the  butyric  fermentation  which 
depends  on  their  presence  ceases  immediately."  (Pasteur.) 

Those  micro-organisms  for  which  free  oxygen  seemed  to  be 
a  poison  were  called  by  Pasteur  anaerobes.  There  exist  "  strict 
anaerobes,"  "  strict  aerobes,"  which  cannot  exist  without  free 
oxygen,  and  "  facultative  "  bacteria  capable  of  living  in  either 
condition. 

In  broth  the  aerobes  form  at  the  surface  a  little  collar,  a 
ring,  or  a  pellicle ;  in  a  drop  of  fluid  under  the  microscope 
they  can  be  seen  to  make  their  way  towards  the  periphery, 
where  there  is  the  best  provision  of  oxygen. 

If  a  filament  from  a  green  alga,  a  plant  containing  chloro- 
phyll, is  put  into  a  suspension  of  motile  aerobes,  and  a  small 
spectrum  of  sunlight  is  allowed  to  fall  upon  it,  the  bacteria  can 
be  seen  collecting  at  the  points  where  the  chlorophyll  assimila- 
tion and  the  production  of  oxygen  are  most  intense,  i.e.,  at  the 
red  and  violet  regions  of  the  spectrum,  the  B  and  C  lines  and 
the  ^line  of  Frauenhofer  (Engelmann's  experiment). 

On  the  contrary,  the  anaerobes  avoid  the  surface  of  the 


PHYSIOLOGY  OF  THE  MICROBES 


81 


drop  of  water  and  the  neighbourhood  of  air  bubbles.  They 
can  be  grown  well  under  shelter  of  a  pellicle  of  aerobic 
bacteria  which  prevent  the  passage  of  air,  this  being  the  best 
method  of  cultivating  the  tetanus  bacillus.  There  is  no 
necessity  to  suppose,  as  does  Kedrowsky,  that  the  aerobes 


d  B  C 


FIG.  35. — Engelmann's  spectrum.  Bacteria  seeking  oxygen  swarming 
round  an  algal  filament  lying  on  a  spectrum.  The  grains  of  chloro- 
phyll are  not  represented.  The  lines  of  the  spectrum  mark  out  the 
regions  on  which  the  bacteria  collect,  i.e.,  the  points  where  most 
oxygen  is  being  liberated. 

secrete  a  special  ferment  which  permits  anaerobic  growth ;  it 
is  sufficient  that  the  anaerobes  are  cut  off  from  free  oxygen. 

The  addition  to  a  tube  of  ordinary  broth,  aerated  and  hence 
unsuitable  for  the  culture  of  anaerobes,  of  sterile  animal  or 
vegetable  tissue,  e.g.,  a  fragment  of  flesh  or  a  piece  of  banana, 
allows  the  anaerobes  to  grow,  the  tissue  acting  as  a  reducing 
agent.  It  is  not  at  all  correct  to  say,  however,  that  the 
anaerobes  live  without  oxygen.  They  only  live,  as  Pasteur 
said,  without  free  oxygen  gas.  They  use  up  the  oxygen  which 
is  present  in  combination  in  the  nutrient  fluid  and  decompose 
the  food-stuffs  to  procure  oxygen  from  them.1 

1  Pasteur  (1861)  :  "There  exist,  besides  the  living  beings  already  known 
which  without  exception,  at  least  in  the  general  opinion,  live  and  breathe 
only  on  condition  of  being  able  to  assimilate  free  oxygen  gas,  others  whose 
respiration  is  so  powerful  that  they  can  live  cut  off  from  the  air  by 

G 


82  MICROBES   AND  TOXINS 

The  decomposition  is  generally  only  partial  and  to  procure 
the  quantity  of  oxygen  and  energy  necessary  the  anaerobes 
have  to  attack  large  quantities  of  the  food-stuffs.  Such 
behaviour  is  typical  of  ferments  and  accordingly  the  anaerobes 
usually  produce  powerful  fermentation.  "  Fermentation  is 
life  without  air,"  was  Pasteur's  dictum.  It  is  in  particular 
the  study  of  alcoholic  fermentation  which  supports  this 
statement.  When  a  yeast  grows  in  a  shallow  mass  of  fluid 
with  an  extensive  surface  its  cells  multiply  abundantly :  there 
is  a  great  increase  in  yeast  protoplasm  but  little  or  no  alcohol. 
When  inoculated  on  the  contrary  at  the  bottom  of  the  fluid 
without  access  of  air,  the  growth  is  feeble  but  produces 
alcohol  in  quantity,  varying  in  proportion  to  the  completeness 
of  the  anaerobic  conditions.  The  differences  in  the  form  and 
functions  of  mucor  when  aerated  or  deeply  immersed  have 
already  been  mentioned.  Several  microscopic  plants,  muce- 
dineae  and  yeasts,  exhibit  a  whole  series  of  transitional  forms 
between  aerobic  and  anaerobic  growths.  Anaerobic  life  appears 
to  be  an  asphyxial  condition  against  which  the  microbe 
contends  or  adapts  itself  by  changing  its  manner  of  nutrition. 
Not  only  the  mucedineae  and  the  yeasts  but  all  living  cells, 
animal  and  vegetable,  act  like  ferments  and  produce  alcohol 
when  forced  to  live  cut  off  from  the  air  in  presence  of  sugar. 
Such  is  the  case  in  the  experiment  of  Pasteur  and  J.  B.  Dumas 
with  the  plums  kept  under  a  bell-jar  :  they  use  up  the  air  and 
fill  the  jar  with  carbonic  acid ;  their  sugar  diminishes  and  they 
become  charged  with  alcohol.  A  similar  case  is  that  of  ripe 
fruits  left  to  themselves  in  an  atmosphere  of  limited  volume, 
as,  for  example,  with  the  apples  and  pears  kept  in  a  closed 
vessel  in  the  experiments  of  Lechartier  and  Bellamy.  Another 
is  that  of  seeds  starting  to  germinate  cut  off  from  oxygen; 
they  produce  alcohol,  using  up  their  reserve  material  (Maze's 

seizing  upon  the  oxygen  of  certain  compounds,  in  which  there  is  in 
consequence  a  slow  progressive  decomposition.  This  group  of  living 
organisms  is  composed  of  ferments  precisely  similar  to  those  of  the  first 
group,  living  like  them,  assimilating  like  them  carbon,  nitrogen,  and 
phosphates,  and  like  them  requiring  oxygen,  but  differing  from  them  in 
their  power  of  doing  without  free  oxygen  gas  and  carrying  on  their 
respiration  with  the  oxygen  derived  from  unstable  compounds." 


PHYSIOLOGY   OF  THE   MICROBES 


83 


experiment).  There  is  even  alcohol  in  animal  tissues.  There 
is,  therefore,  nothing  surprising  in  the  presence  of  alcohol 
throughout  nature,  in  the  soil,  in  water,  in  air,  and  in  the  sea ; 
if  it  is  true  that  the  latter  contains  one  millionth  of  its  weight 
(one  gram  per  cubic  metre)  there  must  be  an  enormous 
supply. 

Since  the  discovery  by  H.  Buchner  of  zymase — the  diastase 
by  which  the  yeast  decomposes  sugar — we  know  that  it  is  on 
the  zymase  rather  than  on  the  anaerobic  conditions  that  the 
alcoholic  fermentation  depends.  But  since  the  zymase  only 
appears  when  the  yeast  is  shut  off  from  the  air,  it  too  is 
"  an  asphyxial  function  "  and  we  return  to  Pasteur's  formula. 

Duclaux  has  re-established  the  continuity  between  the  two 
methods  of  respiration  by  his  idea  of  the  constant  operation 
of  the  zymase  in  aerobic  as  much  as  in  anaerobic  life  and  by 
maintaining  that  alcohol  is  produced  by  living  tissues,  not 
pathologically  but  normally.  "Alcohol  is  a  normal  and 
necessary  product  in  the  digestion  of  the  hydrocarbons  of  the 
seed.  When  oxygen  is  present,  this  alcohol  is  burnt  up  and 
escapes  observation.  To  demonstrate  it  the  plant  must  be 
submitted  to  a  degree 
of  asphyxia  which  just 
lets  it  live,  or  rather, 
which  permits  the  action 
of  the  zymase  which  it 
contains.  It  is  not  the 
asphyxia  which  pro- 
duces the  alcohol,  it 
only  renders  it  per- 
ceptible." 

Further,  absolute 
anaerobiosis  does  not 
exist  either  in  nature  or 
in  our  artificial  cultures.  FlG-  36-— Cochin's  experiment. 

The   pretty   experiment 

of  Denys  Cochin  shows  that  yeast  even  under  anaerobic  con- 
ditions the  most  complete  possible  cannot  do  without  oxygen 

G    2 


84  MICROBES   AND  TOXINS 

indefinitely,  i.e.,  in  traces.  Yeast  cells  are  made  to  grow  shut 
off  from  air  in  a  series  of  communicating  flasks  all  carefully 
sealed.  The  flasks  are  inoculated  in  series,  the  second  with  the 
yeasts  of  the  first  and  so  on.  To  isolate  each  flask  from  the 
preceding  the  little  communicating  tube  is  sealed  in  the  flame. 
Towards  the  tenth  generation,  fermentation  is  seen  to  stop  and 
only  revives  when  a  little  oxygen  is  admitted  to  the  confined 
atmosphere. 

Pasteur  had  already  observed  that  an  air  bubble  about  the 
size  of  a  pin-head  was  sufficient  to  reawaken  a  slackening 
fermentation. 

All  microbes  require  oxygen  but  their  requirements  are  very 
unequal.  Betweeen  the  aerobes  and  the  strict  anaerobes  there 
exists  every  intermediate  condition.  Each  species  requires  an 
oxygen  pressure  suitable  for  itself  just  as  do  the  higher  animals, 
which  are  of  course  aerobic  organisms. 

By  exhausting  the  air  under  a  bell-jar,  Khoudiakow  observed 
that  the  B.  butyricus  could  still  multiply  at  the  pressure  of  five 
millimetres,  Clostridium  butyricum  at  ten  millimetres,  the  vibrion 
septique  and  the  tetanus  bacillus  at  twenty  millimetres  ;  the 
bacillus  of  systematic  anthrax  at  forty  millimetres,  at  which 
pressure  the  latter  microbe  behaves  like  an  aerobe,  using  up 
the  oxygen  in  oxidations. 

An  anaerobe  like  the  B.  butyricus  can  be  trained  to  live  under 
an  oxygen  pressure  greater  and  greater  up  to  fifty  millimetres, 
a  pressure  ten  times  greater  than  that  which  it  will  bear 
normally.  This  acclimatisation  of  anaerobes  to  contact  with 
air  can  be  carried  out  within  certain  limits  in  the  laboratory,  so 
much  so  that  it  has  even  been  thought  useful  to  create  the 
barbarous  phrase  '  aerobisation '  of  anaerobes. 

Khoudiakow  has  made  a  complementary  experiment  by 
modifying  the  pressure  on  aerobes.  B.  subtilis,  grown  on 
gelatin,  lives  fairly  well  under  three  atmospheres,  but  begins  to 
suffer  at  four.  At  the  other  end  of  the  scale  it  still  grows  well 
at  ten  millimetres  of  pressure,  but  not  at  five.  Aspergillus  niger 
has  for  minimum  and  maximum  five  millimetres  and  three 
atmospheres.  Spores  are  more  resistant  to  the  action  of  air 


PHYSIOLOGY   OF   THE   MICROBES          85 

than  bacilli.  The  spores  of  Bact.  butyricum  are  scarcely  affected 
by  the  action  of  air  for  265  days,  whereas  the  growth  of  the 
bacillus  is  inhibited  by  the  action  of  air  for  fifteen  hours. 

Oxygen  is  a  food  which  bacteria  take  from  compounds 
liberating  it  more  or  less  readily ;  anaerobes  take  it  from  com- 
pounds which  retain  it  and  resist  decompositions.  Although  it 
is  not  absolutely  true  that  fermentation  is  life  without  air  (there 
are  fermentations  which  go  on  in  presence  of  oxygen),  it  is  true 
that  anaerobiosis  favours  the  majority  of  fermentations  and  is 
the  usual  condition  for  these. 

In  nature,  anaerobes  occur  wherever  there  is  little  penetra- 
tion of  air,  or  where  the  air  is  diluted  or  replaced  by  other 
gases,  as,  for  example,  in  the  earth,  in  mud  and  slime,  in  sewage, 
in  the  ooze  of  the  sea,  in  dunghills,  and  in  the  intestines  and 
excrements  of  animals  ;  and  it  is  in  these  surroundings  that  the 
most  important  fermentations  and  putrefactions  of  organic 
matter  take  place. 

Respiration  of  the  Pigmented  Bacteria. — The 
purple  bacteria  (a  certain  number  of  which  are  also  sulpho- 
bacteria)  contain  a  ^igmQn^bacterio-purpurine,  quite  distinct  from 
the  pigment  of  Bacillus prodigiosus.  According  to  Engelmann, 
these  pigmented  bacteria  absorb  the  infra-red  rays  of  the 
spectrum  (of  wave-length  0*8  to  0*9  /x)  and  employ  them,  as  also 
the  red  rays,  in  the  decomposition  of  carbonic  acid  from  the 
air  and  the  liberation  of  oxygen,  just  as  plants  do  with  chloro- 
phyll ;  they  have  a  "  chromophyll "  function  analogous  to  the 
chlorophyll  function  of  green  plants.  This  opinion  is  not 
shared  by  all  observers :  according  to  Molisch,  the  purple 
bacteria  are  not  capable  of  decomposing  CO2  nor  of  assimila- 
ting directly  inorganic  compounds.  They  certainly  differ  from 
other  bacteria  in  their  power  of  using  light  in  their  nutritive 
process,  but  their  foods  are  still  organic  food-stuffs  ready  made 
and  they  cannot  do  without  these :  they  are  not  capable  of  the 
synthetic  function  of  the  green  plants.  They  can  assimilate 
organic  food-material  in  the  dark  like  other  bacteria,  but 
they  have  advanced  a  step  by  adapting  themselves  to  light 
and  by  using  it  to  increase  their  nutritive  resources.  But 


86  MICROBES   AND   TOXINS 

they  have  remained  at  that  stage  :  they  have  not  cast  off  the 
necessity  for  ready-formed  organic  food,  i.e.,  the  parasitic  habit, 
nor  can  they  break  up  the  carbonic  acid  of  the  air,  liberating 
oxygen  and  absorbing  carbon.  The  purple  bacteria  then  occupy 
a  position  intermediate  between  the  saprophytic  or  parasitic 
habit  of  bacteria  in  general  and  the  chlorophyll  property  of  the 
higher  plants.  They  carry  on,  like  the  latter,  a  sort  of  photosyn- 
thesis, but  what  they  synthesize  with  the  aid  of  light  is  still 
organic  material  like  the  ordinary  bacteria. 

Secretion  of  Diastases  or  Enzymes. — Microbes  act 
through  their  diastases ;  fermentations  are  thus  diastasic 
reactions.  The  diastases  carry  on  the  transformations  of 
matter  both  by  breaking  down  and  building  up,  and  it  is 
through  them  that  the  bacteria  transform  energy. 

The  discovery  of  diastases  and  the  possibility  of  extracting 
them,  of  isolating  them  (not  completely  pure),  and  of  making 
them  act  without  the  presence  of  living  cells,  represents  a  great 
acquisition  to  the  dominion  of  chemistry  in  the  field  of  the 
study  of  life  and  fermentation. 

A  further  step  was  made  when  Bertrand  demonstrated  the 
prominent  role  taken  by  the  mineral  elements  associated  with 
the  enzymes ;  the  activity  of  laccase  depends  on  the  proportion 
of  manganese  present,  and  the  whole  reaction  behaves  as  if 
laccase  were  a  salt  of  manganese  with  a  weak  acid.  Besides, 
all  the  diastasic  reactions  can  be  performed  by  chemical  agents 
and  the  part  played  by  diastases  seems  to  be  that  of  amplifying 
and  stopping  the  action  of  the  latter. 

The  nature  of  diastases  is  still  unknown,  and  we  shall  not 
dwell  here  upon  the  manner  of  preparing  them,  on  the  causes 
of  error  which  may  creep  into  this  technique,  or  on  the  theory 
of  diastasic  actions  in  general. 

One  microbe  is  capable  of  secreting  several  diastases. 
With  the  proteolytic  enzymes  have  been  grouped  the  lysins 
including  the  haemolysins  of  bacteria.  The  solution  of  the 
cell  attacked  may  indeed  be  only  the  sequel  to  an  action  on 
the  cell  membrane  or  protoplasm,  injurious  but  not  actually 
dissolving.  The  best  known  haemolysins  are  those  of  the 


PHYSIOLOGY   OF  THE   MICROBES          87 

tetanus  bacillus,  tetanolysin;  of  the  staphylococci,  staphylo- 
lysin ;  of  the  cholera  and  pseudo-cholera  vibrios,  vibriolysin ; 
of  the  streptococci,  streptocolysin ;  of  bacillus  pyocyaneus, 
pyocyanolysin ;  lysins  also  exist  in  the  cultures  of  B.  typhosus, 
of  fowl-cholera,  of  the  anthrax  bacillus  and  in  the  bacillus  of 
diphtheria  (in  this  case  doubtless  in  the  body  of  the  bacillus, 
not  excreted). 

The  majority  of  the  bacterial  haemolysins  are  destroyed  by 
heating  to  56°  C. ;  that  of  the  bacillus  of  fowl-cholera,  however, 
is  only  destroyed  at  70°  C. ;  while  pyocyanolysin  stands  boiling 
for  a  long  time  and  is  only  destroyed  in  half  an  hour  at 
120°  C. 

Along  with  these  enzymes  should  be  classed  the  lysins 
which  attack  the  leucocytes  (leucocidine  of  Vandevelde)  and 
other  bacteria  (pyocyanase  of  Emmerich). 

Products  of  Cultures.  Microbic  Excretions. — A 
medium  in  which  a  bacterial  culture  has  grown  contains  bodies 
which  were  not  present  before  the  inoculation.  These  are 
products  of  the  activity  of  the  microbes  in  relation  to  their 
food-stuffs;  they  do  not  come  exclusively  from  the  microbe 
itself,  but  they  are  products  of  these  and  bear  their  mark.  It 
is  often  difficult  to  draw  the  line  between  true  secretory 
products,  the  diastases  and  toxins — and  the  residual  substances 
remaining  after  enzymatic  action,  the  study  of  which  ought  to 
be  included  with  that  of  the  fermentations  and  putrefactions — 
and  the  excreta,  the  catabolic  products,  properly  speaking. 

The  distinctions  which  are  currently  drawn  between  these 
products  depend  often  on  the  purpose  we  have  in  view.  We 
stop  a  fermentation  when  it  has  reached  the  stage  of  the 
products  which  are  useful,  as,  for  example,  in  the  manufacture 
of  beer,  wine,  and  cheese;  if  left  to  continue,  the  organic 
matter  breaks  down  finally  to  the  simplest  substances,  water, 
carbonic  acid,  and  ammonia.  In  nature  a  ferment  only  ceases 
when  it  has  furnished  the  materials  for  a  new  fermentative 
process. 

"A  bacterial  product  is  a  substance  incapable  of  being 
attacked  under  the  conditions  of  the  experiment  by  the 


88  MICROBES   AND   TOXINS 

bacterium  which  has  produced  it,  but  which  can  in  its  turn 
become  a  food  material  if  the  conditions  are  altered,  if  the 
bacterium  takes  on  new  properties,  or  if  other  microbes  step 
in  "  (Duclaux).  Every  living  thing  lives  on  the  products  of 
others. 

As  the  richness  of  a  culture  increases,  its  growth  slackens  : 
the  medium  becomes  less  and  less  favourable,  the  food  material 
becomes  exhausted  and  the  bacterium,  by  no  means  always 
capable  of  living  on  its  own  residues,  ends  by  being  embarrassed 
by  the  substances  it  has  produced.  Acid-producing  bacteria 
cease  to  grow  when  the  acidity  reaches  the  point  where 
vegetation  is  no  longer  possible.  Alcohol  acts  like  an 
antiseptic  towards  the  yeast  that  produced  it  and  acetic 
acid  does  the  same  for  the  ferment  of  vinegar.  The  bacterium 
however  can  often  fall  back  from  the  food  of  its  real  choice  to 
a  sort  of  famine  ration  :  when  the  acetic  ferment  has  used  up 
all  the  alcohol  it  burns  up  the  acetic  acid.  When  a  yeast  has 
no  longer  any  sugar  it  consumes  the  glycerine  which  it  has 
produced  at  its  expense. 

Products  of  excretion  exist  which  stop  the  growth  of  cultures 
by  a  sort  of  auto-intoxication.  The  foulest  waters  are  those 
which  are  least  easy  to  infect,  because,  according  to  Miquel,  they 
contain  substances  of  bacterial  origin  injurious  to  bacterial 
growth.  If  such  foul  waters  are  concentrated  at  a  low 
temperature  and  the  filtered  result  is  added  to  pure  water  this 
latter  becomes  incapable  of  supporting  life.  Boiling  destroys 
these  inhibitory  substances,  which  indicates  perhaps  that  they 
are  of  the  nature  of  diastases.  There  is  said  to  be  in  faecal 
matter  an  inhibitory  substance  which  checks  the  extraordinary 
multiplication  of  bacteria  in  the  intestine,  and  this  also  is  to  be 
regarded  as  a  diastase ;  Conradi  and  Kurpjuweit  compare  its 
energy  to  that  of  carbolic  acid ;  without  having  been  able  to 
isolate  it  they  were  able  by  dialysis  to  make  it  act  without  the 
bacteria  themselves.  They  call  it  an  "  autotoxin."  Others,  how- 
ever, question  this,  not  having  been  able  to  find  it  either  in 
tube  cultures  or  in  the  human  intestine,  and  explain  the 
inhibition  by  the  exhaustion  of  the  medium,  in  the  same  way 


PHYSIOLOGY   OF  THE    MICROBES          89 

that  Pasteur  explained  the  immunity  of  the  body  by  the 
exhaustion  of  the  food  material  which  it  supplies  to  the 
microbe. 

Exhaustion  of  the  food-stuffs  and  action  of  the  excreta  may 
exist  together.  For  example,  when  gelatine  is  inoculated 
with  6  millions  of  bacillus  coli  per  milligram  of  medium 
no  growth  takes  place;  the  bacteria  die  and  disintegrate 
although  the  medium  is  not  exhausted. 

Further,  an  "exhausted"  medium  can  be  regenerated  by 
filtering  it  through  porcelain  and  heating  it  without  adding 
any  new  food  material  (Eijkman).  In  cultures  of  B.  coli  of 
five  days  at  37°  C,  there  is  of  all  the  bacteria  which  can  be 
seen  and  counted  under  the  microscope,  only  one  living  in 
fifteen,  and  after  a  week  only  one  in  forty  (Hehewerth). 
The  antitoxin  of  one  bacterial  species  can  act  on  other 
species. 

Heat  Production. — The  combustion  of  the  food  liberates 
a  quantity  of  energy  which  is  not  entirely  used  up  in  the 
construction  and  support  of  the  bacterial  cells ;  there  remains 
an  excess  of  heat  which  raises  the  temperature  of  the  medium. 
The  yeast  of  beer  undergoing  anaerobic  fermentation  heats 
up  the  whole  mass  by  3*9°  C.  (Eriksson).  In  heaps  of 
manure  or  hay,  the  temperature  may  rise  to  50  or  70°  C., 
and  in  hay  even  to  96°  C.  Cohn  found  in  the  masses  of 
moist  cotton  a  micrococcus  which  discharges  carbonic  acid 
and  raises  the  temperature  to  67°  C.,  when  care  is  taken  to 
avoid  loss  of  heat  by  radiation. 

But  it  is  not  very  certain  that  the  heating  of  hay  is  really 
due  to  microbes.  It  is  only  the  spores  which  can  resist 
temperatures  bordering  on  100°  C.,  and  spores  exert  no 
activity.  Further,  no  bacteria  are  to  be  found  in  the  places 
where  the  heating  begins,  and  finally,  hay  sterilized  at  120°  C. 
can  heat  like  normal  hay. 

According  to  Boekhout  and  Ott  de  Vries,  the  spontaneous 
heating  of  hay  is  a  chemical  phenomenon,  the  cause  of  which 
is  still  unknown. 

Production  of  Light,— Rotting  wood   and  the  corpses 


90  MICROBES   AND  TOXINS 

of  sea-beasts  frequently  emit  light.  Butcher-meat  left  to 
itself  in  a  cool  place  for  two  or  three  days  half-immersed 
in  3  per  cent,  salt  solution  very  often  becomes  luminous. 
Dead  leaves  fallen  in  the  forests  occasionally  give  out  a 
dim,  steady  light.  Now  it  is  not  the  animal  or  vegetable 
tissue  which  shines;  it  must  be  microbes,  moulds,  or 
bacteria. 

Luminous  microbes  have  been  discovered  in  the  Baltic  Sea, 
the  North  Sea  and  the  Indian  Ocean,  and  phosphorescent 
bacteria  in  the  Elbe.  At  present  about  fifteen  moulds  and 
about  thirty  bacteria  are  known  to  be  "photogenic"  or 
"  luminous." 

A  simple  recipe  may  be  given :  take  a  fresh  herring,  sprinkle 
it  with  3  per  cent,  salt  solution,  leave  it  at  a  temperature  of 
about  10°,  add  a  little  sugar,  glycerine,  and  peptone;  in  two 
days  the  flesh  and  the  juice  become  luminous. 

The  production  of  light  depends  on  the  temperature  and  the 
food  supply.  Sometimes  a  temperature  from  20°  to  30°  C. 
suits  best ;  most  often  lower  temperatures  are  more  favourable. 
The  phenomenon  has  been  seen  to  occur  at  +  45°  C.,  and  at 
—  20°  C.  The  luminous  bacteria  seem  to  like  salt  ;  some  only 
require  a  nitrogenous  medium,  others  require  in  addition 
carbon.  But  the  indispensable  substance  is  oxygen,  and  when 
this  is  exhausted  the  luminosity  ceases.  If  a  bubble  of  air  is 
made  to  pass,  by  turning  upside  down,  through  a  long  tube  in 
which  there  is  a  culture  of  a  luminous  bacterium,  which  has 
just  become  extinguished  for  lack  of  oxygen,  a  wave  of  light 
can  be  seen  passing  along  the  tube.  There  is  no  luminosity  in 
a  vacuum. 

Strains  have  been  produced  by  natural  selection  so  luminous 
that  their  light  can  be  seen  in  full  daylight.  If  a  flask  coated 
on  the  inside  with  gelatine  is  inoculated  with  the  Bacterium 
phosphoreum  or  the  Pseudomonas  lucifera,  one  gets  a  microbial 
lamp  which  with  an  eye  a  little  accustomed  to  darkness  allows 
one  to  read  the  time  from  a  watch  or  to  read  moderately  large 
print.  It  might  be  possible  even,  it  seems,  to  employ  such 
lamps  in  powder-magazines  or  mines,  for  they  do  not  emit  heat. 


PHYSIOLOGY   OF  THE   MICROBES          91 

Certain  fishermen  employ  as  bait  luminous  fragments  of  dead 
fish,  a  luminous  bait  provided  by  bacteria. 

With  the  aid  of  the  spectroscope,  Molisch  distinguished  in 
the  light  of  Pseudomonas  ludfera  the  colours  green,  blue  and 
violet.  R.  Dubois  photographed  colonies  of  these  bacteria  with 
their  own  light  alone.  Although  feeble  this  vegetable  luminosity 
exerts,  like  sunlight,  heliotropism ;  the  shoots  of  young  plants 
such  as  vetches,  peas  and  lentils  turn  towards  it  on  germinating  ; 
but  it  is  incapable  of  exciting  the  chlorophyll  function. 

Substances  which  kill  the  microbe  abolish  its  luminosity. 
Dubois  stated  that  he  had  isolated  a  substance  which  shone  on 
contact  with  the  oxygen  of  the  air,  luciferine,  but  others  failed 
to  repeat  this  experiment.  It  is  possible  that  the  microbe 
secretes  a  substance  which  as  soon  as  it  is  produced  is 
destroyed  by  oxidation  giving  out  in  the  process  luminous  rays. 
Many  organic  substances,  aldehydes,  ethereal  oils,  carbides 
of  hydrogen,  fats,  and  alcohols,  when  they  combine  with  free 
oxygen  in  an  alkaline  medium,  can  emit  light  closely  resembling 
that  of  the  above  bacteria;  perhaps  the  bacteria  give  out 
light  thanks  to  the  oxidation  of  substances  in  the  cell,  such 
as  lecithin,  cholesterin,  and  ethereal  fats;  but  there  is  one 
difficulty  :  the  living  cell  does  not  contain  free  oxygen. 

To  prove  that  the  luminosity  of  bacteria  is  a  chemical 
phenomenon  independent  of  the  life  of  the  cell  which  pro- 
duces it,  it  would  be  necessary  to  repeat  with  these  bacteria 
the  experiments  which  have  been  made  with  the  secretions 
of  certain  animal  cells.  With  the  photogenic  substance  of 
Luciola  italica  one  can  write,  and  the  writing  becomes 
luminous  whenever  it  is  moistened.  If  the  luminous  organs 
of  Lampyris  noctiluca,  which  have  been  dried  and  preserved 
in  vacuo,  are  moistened  with  a  drop  of  distilled  water,  the 
luminosity  reappears.  Paper  soaked  in  the  secretion  of 
certain  Myriapods  can  shine  when  moistened,  even  after  two 
months.  In  these  cases  the  luminosity  cannot  be  attributed 
to  a  living  cell. 

Bacteria  which  have  never  been  exposed  to  light  shine  quite 
as  well  as  those  grown  in  daylight.  Their  luminous  property 


92  MICROBES   AND  TOXINS 

is  therefore  not  like  that  of  the  salts  of  strontium  and  barium, 
a  case  of  re-emission  of  light  formerly  absorbed.  It  is  the 
discharge  in  the  form  of  light  of  energy  absorbed  in  another 
form. 

Production  of  Pigments. — Numerous  bacteria  exist, 
whose  cultures  possess  colour,  green,  violet,  red,  blue,  black, 
and  fluorescent ;  these  colours  have  nothing  in  common  with 
the  green  colour  of  chlorophyll  plants,  for  they  are  diffused 
throughout  the  cells,  whereas  chlorophyll  is  agglomerated  in 
distinct  masses.  The  bacteria  which  produce  coloured  cultures 
without  their  cells  themselves  containing  the  pigment  are  the 
more  numerous  ;  the  pigment  is  therefore  an  excretory  product 
which  diffuses  into  the  medium  or  collects  in  little  masses  which 
can  be  seen  under  the  microscope  at  the  side  of  the  bacteria.  All 
the  coloured  bacteria  might  be  put  in  this  category  if  one 
admits  that  the  bacteria  which  contain  a  diffuse  pigment  ought 
to  be  classed  as  algae. 

Staphylococcus  aureus  and  various  sarcinae  produce  colonies 
of  a  golden  yellow :  the  pigment  is  a  fatty  substance  (lipochrome) 
insoluble  in  water  but  soluble  in  alcohol,  benzine,  chloroform, 
ether  and  carbon  bi-sulphide  and  capable  of  saponification  ;  it 
turns  to  blue  or  bluish-green  on  the  addition  of  sulphuric  acid 
and  to  orange  or  red  on  the  addition  of  alkalies. 

Everyone  has  heard  of  the  miracle  of  the  "  bleeding  host" 
the  sacred  bread  which  becomes  covered,  more  by  accident 
than  miracle,  with  red  spots  having  a  reddish-brown,  somewhat 
metallic  lustre.  It  only  means  that  it  has  become  invaded 
by  one  of  the  commonest  bacteria,  one  which  is  present  abund- 
antly in  air,  milk  and  dust,  especially  at  the  end  of  summer  and 
in  autumn,  the  B.  prodigiosus.  The  pigment  is  insoluble  in  water, 
soluble  in  alcohol ;  sulphuric  acid  turns  it  into  reddish-brown, 
alkalies  into  yellow.  Reducing  agents  decolorise  it  as  does 
light,  though  only  after  some  time. 

There  appears  occasionally  on  the  surface  of  milk  a  bluish 
colour,  sometimes  as  a  uniform  film,  sometimes  in  rings  or 
marbling ;  this  is  due  to  the  bacillus  cyanogenes.  The  colour 
is  soluble  in  water,  insoluble  in  alcohol,  ether,  and  chloroform. 


PHYSIOLOGY   OF  THE    MICROBES         95 

Grown  in  pure  culture  in  sterile  milk,  the  colour  is  merely 
grey ;  to  get  the  typical  blue  the  collaboration  of  an  acid- 
producing  bacterium  is  necessary.  In  nature  this  is  provided 
for  by  the  lactic  bacilli.  The  B.  cyanogenes  produces  at  the 
same  time  a  green  fluorescent  pigment. 

The  Bacillus  pyocyaneus  (Gessard)  is  the  colour-producing 
bacterium  which  has  been  most  studied.  It  used  to  be  thought 
that  it  was  the  cause  of  blue  pus  ;  but  it  confined  itself  really — 
in  the  days  before  antiseptics — to  diffusing  its  blue  pigment 
through  the  linen  of  the  dressings.  This  blue  colour,  " pyo- 
cyanin"  is  soluble  in  water  and  chloroform,  insoluble  in 
alcohol ;  it  becomes  pink  in  acid  solution,  yellowish  in  alkaline, 
and  is  a  base  closely  approaching  the  ptomaines. 

The  B.  pyocyaneus  produces  in  addition  a  fluorescent  pigment 
and  a  green  pigment  not  fluorescent ;  and,  finally,  old  cultures 
take  on  a  smoky  brown  tint.  By  heating,  by  inoculating  on 
special  media,  and  by  animal  passages  it  is  possible  to  dis- 
associate or  to  associate  these  different  colours  in  the  same 
microbe  and  to  create  different  strains  or  even  a  non-pigmented 
variety  ;  the  green  fluorescent  pigment  is  particularly  associated 
with  phosphatic  food,  but  the  strains  thus  obtained  depend  on 
the  medium  and  on  the  technique  employed;  they  are  not 
fixed,  and  are  rather  transitory  varieties  than  true  strains. 
The  chromogenic  function  lending  itself  thus  to  modification 
it  is  obvious  that  it  is  not  one  of  the  essential  properties  of  the 
bacterium. 

With  the  microbe,  as  with  higher  creatures,  habits  are  more 
easily  changed  than  nature. 

The  majority  of  the  chromogenic  bacteria  produce  their 
pigment  at  moderate  temperatures,  20  to  25°  C. ;  at  37°  C,  the 
B.  prodigiosus  and  the  sarcina  grow  excellently,  but  produce 
no  pigment.  They  prefer  a  slightly  acid  medium,  but 
fluorescence  requires  the  medium  to  be  alkaline.  The 
starches  are  excellent  food  materials,  which  explains  why  the 
B.  prodigiosus  grows  so  well  on  the  Sacred  Host.  The  essential 
nutritive  material  is  oxygen,  and  with  certain  exceptions  none 
of  them  produce  pigment  when  shut  off  from  air. 


94  MICROBES  AND  TOXINS 

Action  of  Heat  on  Microbes.— Just  as  on  ordinary 
thermometers  the  temperatures  are  marked  for  taking  a  bath 
or  for  keeping  silk-worms,  so  it  would  be  possible  to  mark  the 
points  at  which  microbes  develop  best.  Each  species  has  an 
optimum  temperature;  below  this,  it  grows  feebly;  above  it, 
it  begins  to  suffer  and  dies  j  heat,  indeed,  is  the  sovereign 
disinfectant.  Adapted  as  they  are  to  the  surroundings  which 
shelter  and  nourish  them,  the  bacteria  are  parasites,  not  only 
in  regard  to  their  food  supply,  but  also  for  their  heat 
surroundings. 

Although  the  names  of  different  species  might  thus  be  written 
on  almost  every  degree  of  the  thermometer,  three  types  may  be 
distinguished,  with  intermediate  individuals.  The  majority  of 
the  bacteria  of  water  and  of  soil  and  the  phosphorescent 
bacteria  of  fish  grow  well  at  15 — 20°  C. 

The  majority  of  pathogenic  microbes  demand  in  cultures 
the  same  temperature  as  that  of  the  body  in  which  they  lived 
as  parasites.  The  tubercle  bacillus  of  the  mammals  develops 
best  at  38°  C. ;  that  of  birds  at  41-42°,  and  that  of  fishes  at 
1 5-20°  C.,  /.*.,  practically  like  a  water-bacterium. 

The  third  group  is  that  of  the  thermophilic  bacteria.  They 
demand  and  support  temperatures  so  high  that  other  bacteria 
would  rapidly  be  killed.  They  have  been  found  in  rivers,  in 
sewage,  in  cheese  and  in  the  human  intestine.  The  majority 
are  motile  and  possess  spores.  In  the  hot  springs  of  Ischia, 
and  in  the  fumaroles  of  Naples  there  are  bacteria  which  live  at 
60°  C.  Miquel  found  in  the  Seine  a  species  living  best  at  67° 
to  70°  C.  In  a  spring  at  Luchon,  Certes  and  Garrigou  found 
a  bacterium  developing  at  64°  C.,  the  temperature  of  the  water. 
In  the  upper  layers  of  the  soil,  Globig  discovered  species 
which  grow  well  at  65-70°.  Mile.  Tsiklinsky  has  studied  the 
thermophilic  bacteria  of  the  human  intestine ;  they  are  all 
aerobes. 

It  is  remarkable  that  thermophilic  species  from  the 
surface  of  the  soil  have  been  found  in  the  most  varied 
latitudes,  from  the  tropics  to  the  Hebrides  and  Norway 
Perhaps  those  of  the  cold  countries  (rare)  can. live  shut  off 


PHYSIOLOGY   OF  THE  MICROBES         95 

from  air  at  35-40°  C,  and  can  thus  remain  alive  in  the  intestine 
of  animals. 

The  majority  of  non-sporulating  bacteria  are  killed  in  a  few 
minutes  at  a  temperature  in  the  vicinity  of  60°  C.  Further,  the 
nature  of  the  medium  in  which  they  are  heated  must  be 
reckoned  with  ;  they  perish  more  quickly  in  acid  than  in  alkaline 
fluids,  and  dry  heat  kills  them  much  less  quickly  than  moist. 
The  spores,  being  resistant  forms,  are  only  killed  at  much 
higher  temperatures :  100°  C.,  during  2-4  minutes  for  the 
anthrax  spore.  In  one  single  species,  there  are  spores  which 
stand  the  same  temperature  twice  as  long  as  their  companions. 
At  higher  temperatures  resistance  is  much  shorter,  for  example 
for  certain  sporulating  bacteria  of  the  soil  and  of  hay  (in 
saturated  steam) : 

100°  resistance  ...     .«     16    hours. 

115°          „  .~     ihour. 

130°  »  .«     5    minutes. 

140°          ,, ...      scarcely   I     minute. 

The  spores  of  moulds,  studied  for  the  first  time  by 
Spallanzani,  stand  thirty  minutes  of  dry  heat  at  127-132°  C., 
but  in  moist  surroundings  they  die  below  100°  C.  The 
spores  of  Ustilago  carlo  in  the  presence  of  saturated  water 
vapour  perish  at  about  60°  C. ;  dry,  they  stand  i2o°C.  Spores 
are  more  resistant  than  the  bacilli,  because  they  contain  less 
water,  e.g.,  38  per  cent,  instead  of  62  per  cent.  Tyndall's 
method — discontinuous  heating,  at  intervals,  about  one  hour 
per  day  for  three  days  in  succession — succeeds  at  a  relatively 
low  temperature  because  the  protoplasm  in  taking  up  water 
becomes  more  vulnerable. 

Heating  coagulates  the  protoplasm,  and  this  coagulation  is 
the  more  rapid  and  easy  the  more  water  the  protoplasm 
contains.  Albumin  dried  in  vacua  over  sulphuric  acid  can 
be  heated  beyond  100°  without  losing  its  solubility  in  water 
(Chevreul).  Since  coagulation  is  not  an  instantaneous  but  a 
progressive  phenomenon,  instead  of  talking  of  the  "  tempera- 
ture of  coagulation  "  and  the  "  lethal  temperature  "  it  would 


96  MICROBES   AND  TOXINS 

be  better  to  speak  of  the  lethal  zone  and  of  the  zone  of 
coagulation. 

It  was  from  the  effects  of  heat  on  the  bacilli  that  Pasteur 
discovered  the  anthrax  vaccines. 

Microbes  stand  low  temperatures  very  well.  Long  ago 
Cagniard  de  la  Tour  observed  that  yeast  kept  at  -  90°  C.  in 
a  mixture  of  carbonic  acid  and  ether  does  not  lose  its  power 
of  fermentation.  After  twenty  hours  at  -  130°  C.,  108  hours 
at  -  70°  C.,  the  spores  of  B.  subtilis  still  germinate,  and 
the  spores  of  anthrax  are  still  virulent.  According  to 
MacFadyen's  experiments,  bacteria  kept  for  six  months  at  the 
temperature  of  liquid  air  (about  -  190°  C.)  or  ten  hours  at 
the  temperature  of  liquid  hydrogen  (-252°  C.)  remain  living 
and  virulent. 

Action  of  Light. — Light  is  injurious  to  bacteria  and  is 
thus  a  disinfectant. 

The  active  rays  are  the  chemical  rays  of  the  spectrum  which 
act  by  oxidizing  the  protoplasm  :  the  bacteria  do  not  die  when 
the  sunlight  strikes  them  in  a  vacuum. 

The  anthrax  spores  stand  sunlight  for  about  thirty  hours 
in  contact  with  air  and  eighty  hours  when  shut  off  from  air 
(Roux).  Even  in  vacuo  in  pure  hydrogen  the  bacteria  do  not 
resist  indefinitely.  There  is  therefore  something  else  than 
simple  oxidation  taking  place.  The  action  of  the  air  is 
associated  with  an  action  belonging  more  particularly  to  the 
light,  and  the  oxidation  affects  not  only  the  bacterium  but  the 
medium  in  which  it  is. 

The  bactericidal  rays  are  par  excellence  the  ultra-violet 
rays,  as  can  be  proved  by  cutting  off  certain  parts  of  the 
spectrum  by  means  of  various  sorts  of  screens.  Glass  of  a 
thickness  of  1*35  millimetres  completely  abolishes  the  action. 
A  solution  of  oxalic  acid  of  10  per  cent,  which  limits  the 
spectrum  up  to  300/4^,  acts  in  the  same  way,  whereas  bacteria 
are  destroyed  through  a  screen  of  sulphocyanide  of  potassium 
of  10  per  cent,  which  cuts  down  the  spectrum  to  265/4/4 
(experiments  with  an  electric  arc) ;  the  active  portion  of  the 
rays  of  such  an  arc  must  lie  between  these  limits.  A  blue 


PHYSIOLOGY   OF  THE   MICROBES         97 

specimen  of  rock  salt  cuts  off  all  visible  light  without  cutting 
off  the  active  ultra-violet  portion,  and  rays  which  traverse  it 
destroy  bacteria. 

The  action  of  ultra-violet  rays  is  practically  equally  rapid  in 
the  presence  as  in  the  absence  of  oxygen.  They  produce  a 
little  peroxide  of  hydrogen  in  the  medium  of  suspension,  but 
in  quantities  400  times  too  weak  to  be  active ;  hence  the  action 
is  not  due  to  the  peroxide.  By  putting  in  the  path  of  the 
ultra-violet  rays  from  a  mercury  lamp  a  plate  of  white  glass  of 
one  millimetre  thickness,  all  the  ultra-violet  spectrum  is  cut  off 
beyond  the  rays  3027-3022  ;  the  latter  only  penetrate  the  glass 
very  much  weakened,  and  in  this  case  the  bactericidal  action 
is  much  diminished.  By  far  the  most  powerfully  bactericidal 
rays  are  those  which  have  a  wave-length  below  2*800  units. 
"  Protoplasm  (albumin,  gelatine,  and  serum)  absorbs  the  ultra- 
violet rays  below  2*900  units  :  it  is  therefore  the  rays  absorbed 
by  the  cells  which  exert  the  destructive  action." 

The  ultra-violet  rays  have  been  studied  with  a  view  to  the 
destruction  of  cancer  cells.  Exposed  to  the  ultra-violet  rays  the 
tubercle  bacillus  loses  its  property  of  taking  on  a  stain  which  is 
acid-fast.  An  exposure  of  ten  minutes  kills  them.  An  exposure 
of  one  minute  attenuates  them,  and,  inoculated  in  guinea-pigs, 
they  now  produce  a  slow  lingering  infection ;  the  animal  lives 
for  months,  whereas  the  controls  die  within  forty  days  at  most. 
After  an  exposure  of  three  minutes  the  bacilli  no  longer  grow 
on  potatoes.  The  toxin  of  tubercle,  tuberculin,  which  stands 
heating  at  134°  C.  for  half  an  hour,  is  destroyed  by  five  hours' 
exposure  to  ultra-violet  rays.  The  solutions  should  be  exposed 
n  a  layer  of  two  or  three  millimetres  and  kept  shaken.  Tuber- 
culin exposed  to  the  rays  in  vacua  is  destroyed  much  more 
slowly  than  tuberculin  exposed  in  air  (M.  and  Mme.  Henri 
and  V.  Baroni). 

Certain  coloured  and  fluorescent  substances  such  as  eosin 
erythrosin,  and  bengal-rose,  are  injurious  to  bacteria ;  and  still 
more  so  to  infusoria,  in  presence  of  light,  but  are  quite  or 
almost  harmless  in  the  dark.  These  have  been  called  photo- 
dynamic  substances.  Several  have  been  employed  in  photography 

H 


98  MICROBES   AND   TOXINS 

to  sensitise  plates  towards  rays  which  alone  are  chemically 
inactive.  They  exert  the  same  action  on  ferments  and  on  the 
toxins  and  anti-toxins  of  tetanus  and  diphtheria.  This  action 
is  entirely  due  to  oxidation  and  only  takes  place  when  oxygen 
is  dissolved  in  the  fluids  of  the  experiment.  Thus  in  a  solution 
of  iodide  of  potassium,  with  eosin  added  and  exposed  to  light, 
iodine  is  set  free ;  this  does  not  occur  if  the  solution  is  freed 
from  oxygen.  It  has  been  thought  that  this  oxidizing  action  is 
due  not  to  oxygen  but  to  ozone  (O8). 

Physiology  of  Protozoa. — It  must  not  be  thought  that 
all  the  protozoa  because  they  are  unicellular  are  primitive 
creatures  and  rudimentary  ancestors  of  higher  animals ;  their 
cell  is  adapted  to  all  the  requirements  of  life  and  possesses,  at 
least  in  some  degree,  all  the  properties  of  higher  animals ; 
it  may  be  more  independent,  and  richer  than  certain  cells 
of  vertebrates.  Both  by  structure  and  by  function  the  protozoa 
are  complex  and  highly  differentiated  creatures. 

Ehrenberg,  an  old  scientist,  who  studied  them  very  carefully, 
held  this  belief,  but  in  a  naive  and  inaccurate  form.  Protozoa 
to  him  were  animals  possessing  in  brief  all  the  organs  of 
higher  animals ;  he  saw  in  them  a  digestive  tube,  a  brain,  eyes, 
kidneys,  a  heart,  an  ovary  and  vessels.  But  nothing  of  that 
really  exists ;  the  protozoa  have  simply  nuclei,  vacuoles  which 
digest  the  food,  others  which  expel  the  waste  products  and  a 
protoplasm  full  of  varied  movements  and  currents.  But 
although  they  do  not  possess  the  miniature  organs  which  roused 
Ehrenberg's  admiration,  they  are  none  the  less  capable  of 
taking  up  food,  digesting  it,  and  expelling  the  waste,  of  moving, 
and  of  reproducing.  They  possess  all  the  functions  of  animal 
life,  but  more  simply  and  more  purely  than  among  the  higher 
animals ;  what  one  might  call  the  chemical  and  physical  model 
of  life  is  in  them  more  visible,  more  exposed  to  the  eye. 
That  is  why  their  study  is  so  attractive  and  so  fertile ;  it  is  to 
it  we  owe  our  best  knowledge  and  our  best  ideas  on  life  in 
general,  so  there  is  no  necessity  for  the  surprise  expressed 
by  those  who  have  only  read  their  family  "  Buffon  "  (Buffon's 
Natural  History),  that  scientists  should  be  passionately 


PHYSIOLOGY   OF  THE   MICROBES          99 

studying  infusoria  and  amoebae  rather  than  sharks  and 
elephants. 

Life  means  always  the  combustion  of  protoplasm,  according 
to  Lavoisier's  dictum,  and  this  protoplasm  replaces  its  losses 
by  assimilating  food.  Three  methods  of  assimilation  we  know : 
the  chlorophyllous  plants  with  the  help  of  sunlight  decompose 
the  carbonic  acid  of  the  air,  manufacture  hydrocarbons,  and 
finally  turn  the  starch  into  more  complex  substances  which  go 
to  build  up  their  protoplasm ;  the  green  plant  thus  starts  with 
non-organised  substances  for  all  its  food-supply ;  animals  feed 
on  plants  or  on  other  animals  which  have  already  fed  on 
plants  :  parasites  absorb  food  which  has  already  been  prepared 
ready-made  for  them  by  their  hosts.  All  three  methods 
exist  in  the  protozoa. 

There  are  some  which,  possessing  chlorophyll,  manufacture 
their  food  exactly  like  green  plants  :  these  are  the  plant-animals 
which  form  the  link  between  the  two  kingdoms.  They 
prepare  by  synthesis  a  starch  or  a  para-starch  (Biitschli)  and 
can  satisfy  their  life  conditions  in  water  containing  mineral 
salts,  provided  they  receive  the  light  of  the  sun.  Certain 
species  lose  their  chromatophore  granules  when  ready-made 
food  is  supplied  to  them,  permitting  them  thus  to  dispense 
with  the  labour  of  "  photo-synthesis."  Certain  flagellates  live 
in  symbiosis  with  green  algae  which  supply  them  with  starch : 
in  such  a  case  the  protozoon  may  be  said  to  be  attacked  by  a 
useful  infection :  further,  this  infection  may  be  conveyed  from 
one  to  another. 

The  other  protozoa  capture  their  prey,  frequently  in  the  form 
of  living  creatures.  Whether  they  have  a  mouth  or  not,  whether 
the  food  penetrates  their  bodies  with  the  help  of  a  current 
produced  in  the  water  ^by  cilia  or  flagella,  or  whether 
it  is  the  protoplasm  of  the  protozoon  itself  which  issues 
from  its  envelope,  introduces  itself  into  the  body  of  the 
prey,  and  thus  devours  it  from  the  inside,  as  it  were  re- 
versing the  roles,  in  all  cases  the  important  point  in  the 
nutrition  is  intracellular  digestion :  the  food  is  enclosed  in  a 
little  spherule  inside  the  protoplasm  where  little  by  little  it  is 

H  2 


100 


MICROBES  AND  TOXINS 


\ 


\vvW 

\\\1  IX  i  Ml 


FlG.  37. — Gromia  oviformis  in  the  act  of 
capturing  in  its  pseudopodia  a  diatom, 
which  being  too  large  for  ingestion  is 
digested  outside  the  body  in  this  way. 
(After  Max  Schultze.) 


dissolved  by  means  of 
the  digestive  juices. 
A  leucocyte  which 
seizes  and  digests  a 
bacterium  in  a  higher 
animal  acts  in  precisely 
the  same  way.  The 
amoeba  is  the  proto- 
type of  that  phagocytic 
digestion  which  occu- 
pies so  large  a  place  in 
both  natural  and  ac- 
quired immunity. 

Of  what  nature  is 
this  digestion  from  the 
point  of  view  of  chem- 
istry ?  In  the  digestive 
vacuoles  the  reaction 
is  acid  and  from  certain 
myxomycetes  (Fuligo 
varians)  and  rhizopods 
(Pelomyxa  palustris) 
a  ferment  resembling 
pepsine  and  acting  in 
an  acid  medium  has 
been  isolated.  On  the 
other  hand,  Mouton 
and  Mesnil  have  ex- 
tracted from  amoebae 
and  paramecia  a  fer- 
ment which  digests 
gelatine  and  fibrin  in 
an  alkaline  medium 
just  like  trypsin. 

According  to  other 
investigators  the  diges- 
tive medium  is  first 


PHYSIOLOGY   OF  THE   MICROBES        101 

acid,  then  alkaline,  just  as  in  the  stomach  and  small  intestine 
of  mammals. 

The  solid  refuse  of  digestion  is  evacuated  by  an  anus  and  the 
liquid  residue  collects  inside  protoplasm  in  a  little  spherical 
sac    which     from 
time   to   time  ex- 
pels  its    contents 
externally :     this 
latter  is  the  con- 
tractile     vesicle 

which    Ehrenberg 

,     f        .  .         FIG.  38. — An  amoeba  expelling  the  residue  of  its 

took  for  the   pul-  food :  various  stages.     (After  Verwom.) 

sating  heart. 

Oxygen  is  a  primary  necessity  to  protozoa  as  to  bacteria  :  the 
digestive  vacuoles  contain  oxygen,  the  contractile  vesicles 
discharge  carbonic  acid,  i.e.,  aerobic  respiration.  The  protozoa 
which  live  in  surroundings  deprived  of  free  oxygen  have,  it  is 
certain,  a  method  of  respiration  analogous  to  that  of  anaerobic 
bacteria ;  they  draw  their  oxygen  from  reserve  materials  which 
they  have  stored  within  themselves,  e.g.,  glycogen.  It  is 
believed,  without  being  absolutely  certain,  that  certain  infusoria 
can,  like  the  intestinal  worms  (ascarides),  break  up  glycogen 
with  the  formation  of  valerianic  and  carbonic  acids.  Among 
the  products  of  excretion  have  been  found  uric  acid  and 
phosphate  of  lime  (Schewiakoff).  Excretion  is  a  fairly  active 
process  since  the  vacuole  contracts  often  (every  four  to  eighteen 
seconds  according  to  temperature  in  Stylonychia  pustulata). 
According  to  Maupas  the  infusoria  discharge  during  a  space 
of  time  which  varies  from  two  to  forty-six  minutes  a  volume  of 
liquid  equivalent  to  the  whole  volume  of  the  animal. 
Stimulation  from  the  exterior  is  always  accompanied  in  the 
protozoa  by  a  manifestation  of  energy :  they  possess  irritability. 
The  excitant  may  be  a  touch,  a  ray  of  light,  heat,  electricity  or 
a  chemical  substance,  and  the  protozoon  in  its  reactions  to  the 
stimulus  acquires  habits  such  that  its  physiology  is  full  of  as 
many  problems  as  that  of  the  higher  animals,  not  excepting 
problems  in  psychology.  Although  possessing  neither  nervous 


102  MICROBES   AND  TOXINS 

system  nor  sense  organs,  even  to  such  a  degree  as  the  sea- 
anemone,  the  protozoon  is  capable  of  choice  and  of  determina- 
tion, these  phenomena  of  course  remaining  more  or  less 
mechanical.  Superposition  and  propagation  of  impressions 
exist  among  protozoa,  and  after  long  and  minute  observations 
it  has  been  maintained  that  no  essential  difference  exists 
between  them  and  the  most  complicated  metazoa :  activity  is 
neither  more  nor  less  mechanical  in  the  one  set  than  in  the 
other. 

The  chief  business  of  all  living  creatures  is  reproduction. 
Among  protozoa  there  exist  several  principal  methods  for  this, 
each  presenting  numerous  variations.  They  may  divide  by 
nuclear  division  :  they  may  divide  by  budding,  and  in  this  the 
greatest  diversity  occurs  in  the  number,  size,  and  arrangement 
of  the  buds.  They  may  sporulate,  /.*.,  their  protoplasm  may 
break  up,  and  each  fragment  consisting  of  a  bit  of  protoplasm 
and  a  bit  of  nuclear  material  can  reproduce  a  creature  similar 
to  the  mother-cell  which  sporulated.  When  life-conditions 
are  difficult,  certain  protozoa  encyst,  *.*.,  they  contract  inside 
a  resistant  shell,  and  under  shelter  of  this  various  modes  of 
reproduction  may  take  place.  Between  reproduction  by 
division  and  that  by  sporulation  intermediate  forms  exist 
(Tillina  and  Colpidtum\  and  the  continuity  in  nature  can 
always  be  detected  by  the  imagination. 

Parasitic  protozoa,  those  which  to  subsist  require  to  emigrate 
from  one  host  to  another,  most  often  reproduce  themselves 
by  sporulation,  and  the  reproduction  is  the  more  lavish  the 
greater  the  difficulties  encountered  by  the  species  in  propa- 
gation. 

Parasitism  tends  to  modify  the  species  in  a  retrograde 
direction  but  the  losses  may  be  compensated  for  by  new 
acquisitions.  Locomotory  organs,  protective  envelopes  and 
the  apparatus  designed  for  capturing  and  digesting  food 
become  simplified  or  disappear  altogether.  But  parasites  are 
in  general  more  prolific  and  they  acquire  other  organs,  hooks 
or  suckers,  by  which  they  can  better  cling  to  their  host.  As 
their  life  conditions  or  their  habits  become  narrowed  down, 


PHYSIOLOGY   OF  THE    MICROBES        103 


they  present  those  phenomena  of  strict  adaptation  which  are 
equivalent  as  between  the  soil  or  the  host  and  the  parasite — to 


formation  and  dischatye 
oTUt&  sporozolles 


a  sporvzoite  invading  ffus 
epithelial  cell  andvecomina 
adult 


division  of 
nucleus 


FlG.  39. — Life  cycle  of  a  Coccidium  (Coccidium  Schubergi). 
(After  Schaudinn.) 

true  specificities :  they  cannot  endure  any  other  habitat,  food, 
or  host. 

For  example ,  Cost 'i :a   Jiecatrix  which  lives  attached    to    the 


104  MICROBES   AND   TOXINS 

surface  of  the  bodies  of  fishes  can  no  longer  live  even  in  the 
same  water  when  it  becomes  detached  and  floats  free.  The 
infusoria  from  the  paunch  of  ruminants  or  the  caecum  of 
horses  cannot  live  outside  the  bodies  of  these  animals  except 
at  body  temperature,  37°C. :  they  are  thus  examples  of 
semi-parasitism.  The  parasites  of  mammalian  blood  are 
habituated  or  even  confined  to  life  at  definite  temperatures ; 
hence  the  effects  of  climate  and  season. 

The  same  parasites  attach  themselves  to  one  host  only  and 
their  presence  becomes  a  specific  character  of  the  latter.  The 
parasite  of  malaria  is  peculiar  to  man  and  among  the  mosquitoes 
it  can  only  inhabit  those  of  the  genus  Anopheles.  But  one 
trypanosome  can  live  in  several  species  of  host,  one  species 
often  serving  as  a  sort  of  reservoir  for  others  (it  is  probable,  for 
example,  that  cattle  form  a  reservoir  for  the  Trypanosoma 
gambiense,  which  causes  sleeping  sickness  in  man). 

Lamblia  intestinalis  is  a  parasite  of  the  small  intestine : 
gregarines  are  only  found  in  the  large  intestine,  in  the 
peritoneum,  and  in  the  genital  organs  of  their  hosts  (inverte- 
brates) :  coccidia  inhabit  the  epithelial  cells  :  the  haemosporidia 
of  malaria  only  the  red  corpuscles  of  the  blood :  while  the 
sarcosporidia  only  occupy  the  muscle  cells.  But  parasites 
exist  which  infect  all  the  organs  of  the  host,  e.g.,  Myxobolus 
pfeifferi  in  the  barbel  disease. 

Chemiotactic  phenomena,  positive  or  negative,  are  observed 
among  protozoa  as  among  bacteria,  and  it  is  by  an  action  of 
this  sort  or  by  a  choice  of  soil  (which  closely  resembles  it) 
that  the  affinity  of  the  sporozoites  of  the  malarial  haemo- 
sporidia  is  explained  for  the  salivary  glands  of  the  mosquito 
which  inoculates  man.  The  hgemosporidia  sucked  from  the 
blood  of  the  patient  gain  the  stomach  of  the  mosquito  and 
there  enter  upon  the  sexual  cycle,  a  cycle  which  cannot  go  on 
in  any  other  surroundings  :  this  specific  action  is  no  doubt  due 
to  certain  physical  and  chemical  conditions  which  are  only 
realised  there  and  of  which  the  following  fact  may  give  some 
idea :  the  appearance  of  sexual  forms  in  human  blood  is 
favoured  by  adding  a  little  distilled  water  to  the  blood  of 


PHYSIOLOGY   OF  THE   MICROBES       105 

a  preparation  (Manson).  The  ideas  of  specificity  "  of  soil  " 
and  of  virulence  must  resolve  themselves  among  the 
protozoa  as  among  the  bacteria  into  physical  and  chemical 
factors. 

The  parasitic  protozoa  of  the  intestine  of  one  host  pass 
into  another  host  through  the  external  world  in  the  state  of 
spores  or  cysts.  The  parasites  of  the  blood  cannot  enter 
the  blood  of  a  new  individual  (in  nature)  except  through  the 
intermediation  of  a  blood-sucking  insect,  and  a  portion  of 
their  life-cycle  takes  place  in  this  intermediary.  Thus  the 
parasite  of  Laveran  is  inoculated  from  man  to  man  by  the 
Anopheles  mosquito.  In  these  cases  the  principal  host  is  the 
one  in  which  the  sexual  phase  of  the  life-cycle  takes  place ; 
the  host  in  which  occurs  the  non-sexual  reproduction  is  only 
the  secondary  or  "intermediate  host."  With  regard  to  the 
parasite  of  malaria  man  is  the  intermediate  or  secondary  host, 
and  the  principal  host  is  the  mosquito.  The  tsetse  fly  is  the 
principal  host  of  the  Trypanosoma  gambiense  of  sleeping 
sickness. 

Though  the  protozoa  are  frequently  parasites,  they  are  often 
themselves  attacked  by  parasites,  by  bacteria,  by  chytridiaceae, 
by  saprolegnaceae,  and  by  algae.  The  algae,  however,  may  be 
useful  commensals,  furnishing  a  food-stuff — starch ;  but  other 
parasites  may  kill  the  protozoa  which  they  infect,  provided 
the  latter  does  not  defend  itself  and  overcome  its  parasite 
by  devouring  and  digesting  it — again  by  intracellular 
digestion. 

To  study  the  bacteria  pure  cultures  can  be  made  in  which 
they  find  their  food  material  in  solution.  The  protozoa  have, 
doubtless,  methods  of  nutrition  much  more  complicated,  for 
their  culture  is  more  difficult.  Several  trypanosomes,  parasitic 
in  the  blood,  or,  more  precisely,  in  the  blood  plasma,  have, 
been  grown  in  pure  culture  by  Novy  and  MacNeal  on  media 
to  which  blood  was  added.  The  parasites  of  malaria  (parasites 
of  the  red  corpuscles)  have  not  been  cultivated.  The  amoebae 
can  be  grown  on  condition  that  suitable  prey  is  supplied  :  if 
in  a  culture  there  are  only  amoebae  and  as  prey  a  bacterium 


106  MICROBES   AND   TOXINS 

for  example  the  B.  coli  in  pure  culture,  the  culture  is  said 
to  be  "  pure-mixed."  The  prey  may  consist  of  a  dead 
bacterium. 

In  the  absence  of  cultures  experiment  is  difficult ;  the  study 
of  protozoa  is  still  necessarily  attached  to  the  study  of  their 
forms,  and  physiological  study  takes  of  necessity  the  second 
place  to  morphological.  The  method  par  excellence  consists 
in  following  the  life-cycle  of  protozoa  in  their  natural 
surroundings  or  in  their  hosts.  The  study  of  bacteria  has 
been  capable  of  greater  advances,  thanks  to  pure  cultures  in 
well-defined  media  which  permit  of  chemical  analysis.  The 
application  of  similar  methods  to  protozoa  is  infinitely 
desirable,  not  only  for  the  sake  of  medicine,  but  also  to  extend 
our  knowledge  of  the  phenomena  of  life  in  general. 


CHAPTER  V 

PATHOGENIC  MICROBES — INFECTION 

ORIGIN. — Specificity — Virulence. — How  virulence  may  have  been  acquired 
— Evolution  of  microbes — The  '  para  '  and  the  '  pseudo  '  forms — 
Diminution  and  augmentation  of  virulence — Pasteur  and  attenuation  of 
the  virus. 

INFECTION. — The  conflict  between  the  microbe  and  the  body — Methods 
— of  transmission — Latent  microbism — Germ-carriers — The  number  of 
microbes  sufficient  to  produce  infection — Microbial  associations— Paths 
of  penetration  and  inoculation — The  role  of  the  intestine — Seats  of 
election  and  susceptible  cells — Incubation. 

ORIGIN  :  SPECIFICITY  :  VIRULENCE. 

THE  idea  of  pathogenic  microbes  arose  as  a  result  of  Pasteur's 
labours  on  fermentation. 

For  years  the  bacteridium  had  been  seen  in  the  blood  of 
anthrax  animals  without  giving  rise  to  the  thought  that  these 
microscopical  rods  were  the  cause  of  the  illness.  After  the 
discovery  of  the  bacillus  of  butyric  fermentation,  Davaine 
considered  that  the  bacteridia  were  the  cause  of  anthrax  as  a 
sort  of  peculiar  fermentation  having  for  its  subject  the  body  of 
an  animal. 

We  do  not  yet  know  the  pathogenic  microbes  though  they 
certainly  exist  in  small-pox,  in  vaccinia,  in  measles,  in 
scarlatina,  in  mumps,  and  in  hydrophobia.  That  of  syphilis 
remained  unknown  up  to  1905.  Pasteur  was  the  first  to  handle 
invisible  microbes  with  sufficient  confidence  to  discover -a 
method  of  vaccination.  His  discoveries  in  hydrophobia  aroused 
researches  on  the  so-called  invisible  microbes.  A  nervous 

107 


108  .    MICROBES   AND   TOXINS 

disease  which  resembles  hydrophobia,  the  acute  poliomyelitis 
of  children,  has  recently  been  studied  by  the  same  experimental 
method. 

The  microbial  doctrine  has  still  opponents,  more  or  less 
masked,  who  accuse  microbiologists  of  being  able  to  see 
nothing  but  the  microbe  and  of  imagining  that  this  is  the 
whole  malady.  The  body  takes  some  part,  there  is  no  doubt ; 
the  malady  is  a  sort  of  fermentation,  but  one  taking  place  in  a 
medium  capable  of  resisting  the  ferment.  Pasteur  recreated 
medicine  by  introducing  into  it  the  spirit  and  method  of  the 
exact  sciences,  but  he  knew  as  well  as  any  that  diseases  do  not 
rage  in  an  inert  material.  This,  however,  does  not  prevent  the 
various  incidents  of  the  disease  from  being  at  bottom  physico- 
chemical  phenomena. 

The  Origin  of  Pathogenic  Microbes. — The  pathogenic 
microbes  are  not  instruments  of  a  perfidious  Providence,  and 
created  to  chastise  man,  animals,  and  plants.  The  pathogenic 
species,  are  species  the  result  of  selection  and  adaptation.  They 
grew  first  of  all  as  saprophytes  on  individuals  who  suffered  no 
damage,  as  is  the  case  to-day  with  many  bacteria  growing  on 
animal  bodies.  They  multiplied  upon  ill-nourished  and 
fatigued  individuals  and  found  on  a  definite  animal  species 
nutritive  materials  and  a  chemical  "  soil "  which  suited  them. 
Certain  bacteria  have  become  strict  parasites,  incapable  of 
living  even  temporarily  in  the  external  world,  e.g.,  the  bacillus  of 
leprosy.  These  views  of  Pasteur  are  quite  in  conformity  with 
the  spirit  of  Darwinism. 

From  the  beginning  there  has  been  happening  what  occurs 
every  day,  i.e.,  there  has  been  a  struggle  between  the  parasite 
and  the  body.  Not  only  does  the  body  defend  itself  against 
the  bacteria,  but  the  bacteria  defend  themselves  against  the 
body.  Each  is  capable  of  gathering  strength  or  immunising 
itself  against  the  other,  and  these  are  simply  different  aspects 
of  adaptation  and  of  natural  selection.  "  The  science  of 
bacteria,  as  with  all  the  branches  of  biology,  has  profited  by 
the  theory  of  evolution  and,  making  a  just  return,  it  has 
supplied  the  Darwinian  theory  with  a  striking  confirmation. 


PATHOGENIC   MICROBES— INFECTION     109 

The  great  discovery  of  Pasteur  of  the  attenuation  of  viruses 
proves  the  plasticity  of  microbial  species  and  the  facility  with 
which  they  modify  their  primitive  characters.  The  history  of 
bacterial  diseases  shows  also  the  great  role  which  these  in- 
finitesimal creatures  have  played  in  natural  selection,  for  is  it 
not  they  which  have  caused  to  disappear  in  the  course  of  ages 
certain  vegetable  and  animal  species  insufficiently  armed  to 
resist  them?  .  .  .  Experimental  medicine  has  studied  the 
adaptation  of  certain  pathogenic  microbes  which  permit  them 
to  attack  the  body  in  spite  of  the  defences  opposed  to  them. 
Here  it  is  probably  a  question  of  a  selection  of  individuals 
endowed  with  particularly  stable  characters." l 

Starting  with  a  bacterium  almost  non-virulent,  Pasteur 
succeeded  in  infecting  with  anthrax  in  succession,  by  the 
method  of  passages,  the  new-born  mouse,  the  adult  mouse,  the 
young  guinea-pig,  the  adult  guinea-pig,  the  rabbit,  and  the 
sheep.  Vincent  by  introducing  into  the  peritoneal  cavity  little 
collodion  sacs  containing  bacteria  which  are  nourished  by  the 
body  fluids,  while  being  protected  from  the  cellular  defences, 
rendered  pathogenic  for  the  guinea-pig  and  the  rabbit  such 
saprophytic  microbes  as  B.  megatherium  and  B.  mesentericus 
vulgatus,  but  the  virulence  thus  acquired  disappeared  as  soon 
as  the  artifice  which  produced  it  was  suspended.  These 
experiments  do  not  permit  of  the  conclusion  that  pathogenic 
species  can  be  created  at  will  in  the  laboratory ;  only  more  or 
less  stable  variations  are  got  It  is  not  with  such  ease  that  we 
are  likely  to  reproduce  what  Nature  has  taken  centuries  to 
accomplish.  It  is  very  probable  that  small-pox  and  vaccinia 
are  two  modifications  of  the  same  virus.  Yet  the  production 
of  vaccinia  with  small-pox  has  not  yet  been  successfully 
performed.  The  experiments  said  to  have  proved  variolo- 
vaccination  are  still  disputed.  Nothing  has  been  able  to 
produce  from  B.  coli  a  Typhoid  bacillus.  There  exist  various 
families  of  the  Tubercle  bacillus,  which  may  be  secular  adapta- 
tions from  the  same  strain,  adaptations  to  the  human  species 

1  Metchnikoff,  address  read  to  the  Cambridge  festival  in  honour  of  the 
Darwin  centenary,  June,  1909. 


110  MICROBES  AND  TOXINS 

and  to  the  ox,  to  birds,  reptiles,  frogs,  and  fishes,  but  no  one 
has  ever  succeeded  in  producing  a  tubercle  bacillus  of  the 
human  type  from  the  tubercle  bacilli  (or  acid  fast  bacilli)  of 
the  frog.  Nocard's  experiments  in  which  he  transformed  the 
human  bacillus  into  the  bird  bacillus  by  repeated  passages 
have  not  been  confirmed.  The  researches  of  recent  years 
confirm  the  idea  of  Th.  Smith  and  R.  Koch,  that  the  human 
bacillus  and  the  bovine  bacillus  cannot  be  transformed  one 
into  the  other,  at  least  under  the  time  conditions  of  our 
experiments ;  the  fixity  of  these  acquired  characters  has  even 
raised  the  hope  that  it  might  be  possible  to  vaccinate  cattle 
against  bovine  tuberculosis  by  means  of  bacilli  of  the  human 
type  and  vice-versd,  perhaps,  men  with  the  bovine  bacillus  or 
products  derived  from  it. 

The  Darwinian  conception  of  evolution  in  pathogenic  mi- 
crobes is  nevertheless  true,  although  direct  proofs  are  lacking. 
Similarly  the  simian  origin  of  man  would  be  quite  as  certain, 
although  some  of  the  proofs  were  lacking,  and  even  although 
we  might  not  be  able  to  demonstrate  forms  intermediate 
between  man  and  monkey. 

Specificity. — It  is  necessary  to  distinguish  between  the 
specificity  of  the  microbe  and  that  of  the  disease. 

Typhoid  fever  is  caused  by  the  bacillus  of  Eberth  and  by 
it  alone — specificity  of  the  disease.  The  typhoid  bacillus 
remains  the  typhoid  bacillus  in  cultures  and  in  the  intestine ; 
it  does  not  tend  towards  either  the  dysentery  bacillus  or  the 
coli  bacillus — specificity  of  the  microbe.  These  two  ideas  hang 
together ;  the  same  causes  must  produce  the  same  effects  if 
there  is  to  be  any  science  at  all,  but  to  produce  the  same  effect 
it  is  absolutely  necessary  that  the  cause  remain  the  same. 

A  disease  appears  with  certain  symptoms  and  anatomical 
lesions  due  to  a  definite  microbe,  but  these  symptoms  and 
these  lesions  may  be  produced  by  others  ;  for  example,  the 
tubercle  is  the  lesion  par  excellence  produced  by  the  tubercle 
bacillus  of  Koch,  but  tubercles  are  also  produced  by  the 
bacillus  of  glanders.  There  are  not  so  many  possibilities  of 
reaction  in  the  body  as  there  are  bacteria;  fever,  effusions, 


PATHOGENIC   MICROBES— INFECTION     111 

congestion,  false  membranes,  tubercles,  are  all  properties  of 
the  body  rather  than  of  the  microbes.  The  specificity  of  the 
disease  consists  in  a  definite  combination  of  symptoms  along 
with  the  invariable  presence  of  a  definite  bacterium.  To 
render  possible  the  study  of  disease  it  was  a  primary  necessity 
for  a  certain  disease  to  be  the  result  of  a  certain  micro- 
organism, and  further  for  this  latter  to  be  of  fairly  stable 
natural  characters.  The  typhoid  bacillus  causes  typhoid  fever, 
but  if  it  were  capable  of  changing  its  characters  the  hygiene 
and  prophylaxis  of  this  disease  would  be  without  any  sure 
foundation. 

Microbiology,  medicine,  and  hygiene  therefore  can  no  more 
do  without  this  idea  of  specificity  than  science  in  general  could 
exist  without  the  idea  of  causality. 

Medicine  has  always  been  pursuing  this  conception,  but  has 
only  finally  seized  it  by  the  help  of  microbiology  and  chemistry. 
Even  the  beliefs  in  "  miasmata  "  and  "  epidemic  causes  "  were 
already  attempts  in  this  direction.  Common  sense  has  always 
been  a  believer  in  the  specificity  of  transmissible  diseases.  It 
was  for  that  reason,  as  we  read  in  the  Old  Testament,  that  the 
Israelites  isolated  lepers.  Herodotus  knew  that  leprosy  passes 
from  man  to  man ;  Galen  believed  in  the  specificity  of  hydro- 
phobia, of  scabies,  of  granular  conjunctivitis.  The  idea  of 
a  specific  disease  was  bound  to  suggest  the  idea  of  a  specific 
agent. 

The  fact  that  those  who  have  had  small-pox  scarcely  ever 
take  it  a  second  time,  but  are  still  quite  virgin  soil  for  measles 
and  scarlatina  and  vice  versa,  spoke  again  in  favour  of  the 
specificity  of  diseases.  Jenner's  discovery  even  furnished 
a  general  principle  of  diagnosis  between  specific  viruses. 

In  favus,  in  pityriasis  versicolor,  in  the  ring-worms,  and  in 
thrush,  there  had  already  been  observed  before  the  days  of 
Pasteur  the  constant  presence  of  the  same  microscopical  fungi, 
and  it  had  even  been  concluded  that  these  diseases  were  due 
to  parasites.  But  the  opponents  of  this  idea  maintained 
(there  are  perhaps  still  existing  some  who  believe  this)  that 
these  organisms  were  not  the  cause,  but  in  a  way  constant 


112  MICROBES   AND  TOXINS 

"witnesses"  of  the  diseases,  and  simply  the  inhabitants  of 
lesions  which  they  had  not  produced — just  as  the  same  moulds 
are  usually  found  in  pots  of  the  same  jam  left  open  to  the  air. 

The  specificity  of  infectious  diseases  has  been  demonstrated 
by  Pasteur  and  Koch. 

Since  there  is  no  "  spontaneous  generation,"  at  least  in  the 
world  of  the  present  day,  it  is  impossible  for  the  micro- 
organisms to  originate  in  the  diseased  tissues,  and  the  science 
of  fermentation  has  proved  that  a  given  cell,  inoculated  in  a 
sterile  fluid  of  known  composition,  produces  in  it  definite  and 
constant  phenomena.  These  ideas  were  taken  up  by  medicine 
when  Davaine  maintained  that  the  bacteridium  was  the 
cause  of  anthrax,  and  when  Obermeier  held  that  recurrent 
fever  was  caused  by  the  spirochaete  found  in  the  blood  of  the 
patients  during  the  fever. 

To  prove  the  specific  activity  of  a  ferment  it  is  necessary  to 
isolate  and  make  pure  cultures  of  it  and  to  re-inoculate  it.  It 
was  this  that  we  learnt  from  Pasteur  and  Koch.  Koch's 
memorandum  on  tuberculosis  remains  the  complete  model 
for  the  discovery  of  the  microbe  of  a  disease  and  of  the 
demonstration  of  its  specificity. 

Specificity  of  function  is  the  point  of  capital  importance. 
The  fixity  of  form  in  bacteria  is  of  great  use  in  the  search  for 
and  identification  of  them ;  but  on  this  lacter  point  science  has 
been  obliged  to  become  less  exacting ;  the  form  of  the  bacteria  is 
not  always  a  sufficient  distinguishing  character.  They  are 
really  defined  by  their  chemical  and  physiological  actions. 
Thus  the  tubercle  bacillus  is  better  defined  by  its  staining 
peculiarities  (a  physico-chemical  reaction)  than  by  its  shape; 
still  better  by  the  appearance  of  pure  cultures  than  by  staining; 
and  better  still  than  by  the  cultures,  by  its  excretion  of 
tuberculin.  Finally,  the  study  of  the  reactions  of  the  body, 
*.*.,  of  immunity,  has  shown  that  the  cells  of  the  patient  respond 
in  a  specific  manner  to  the  attack  of  the  bacterial  cells,  and  both 
medicine  and  hygiene  daily  employ  the  property  of  specificity 
in  the  antibodies. 

The  "  Para  "  and  "  Pseudo  "  Bacteria.— We  may  now 


PATHOGENIC   MICROBES— INFECTION     113 

go  on  to  cite  a  series  of  facts  which  have  modified  the  idea  of 
rigorous  specificity  among  microbes,  a  specificity  which  in  the 
early  days  of  medical  bacteriology  was  believed  to  be 
absolute ;  or  at  least  these  facts  have  restricted  this  idea  (by 
compelling  us  to  create  new  varieties),  although  we  may  still 
regard  it  as  sufficient  for  the  purposes  of  medicine  and  hygiene. 

It  was  long  the  custom  to  talk  simply  of  the  bacillus  of 
diphtheria,  the  typhoid  bacillus,  the  B.  of  dysentery,  the 
meningococcus,  the  cholera  vibrio,  etc.,  but  little  by  little 
there  have  been  discovered  bacteria,  close  relatives  of  each  of 
these  typical  microbes,  but  not  possessing  all  their  characters. 

From  the  time  of  the  first  bacteriological  discoveries  in 
diphtheria,  bacilli  were  isolated  from  the  mouth  exactly 
similar  to  the  pathogenic  bacillus,  but  non-toxic ;  the  best 
known  is  that  described  under  the  name  of  Hoffmann's 
bacillus.  They  have  been  called  pseudo-diphtheria  bacilli,  and 
have  been  found  in  diphtheritic  sore  throat,  in  scarlatinous  sore 
throat,  and  in  the  normal  conjunctiva,  even  sometimes  in 
vaccine  lymphs.  Some  of  them  are  pathogenic  for  the  guinea 
pig,  and  produce  in  it  a  septicaemia  bearing  no  resemblance  to 
diphtheria.  They  are  not  affected  by  antidiphtheritic  serum 
and  are  incapable  themselves  of  being  used  to  produce  such 
a  serum.  Roux's  opinion  was  that  it  was  a  question  of 
degenerated  diphtheria  bacilli,  or  of  bacilli  not  yet  adapted, 
not  having  yet  found  the  conditions  capable  of  exalting  their 
virulence.  It  must  be  added  that  the  name  of  pseudo- 
diphtheria  has  been  incorrectly  applied  to  bacilli  which  do 
not  deserve  the  name,  even  by  their  form. 

It  is  no  use  playing  with  words.  It  is  on  the  clinical  facts 
the  problem  ought  to  depend.  There  are  found  in  the  most 
typical  cases  of  diphtheritic  sore  throat  diphtheria  bacilli 
possessing  every  degree  of  toxicity,  and  also  bacilli  which  are 
not  pathogenic  (i.e.,  for  the  guinea-pig,  since  they  cannot  be 
inoculated  in  man).  All  these  bacilli  are  called  diphtheria 
bacilli.  On  the  other  hand,  there  exist  non-diphtheritic 
affections  both  of  the  throat  and  of  the  nose  in  healthy 
individuals  where  bacilli  resembling  diphtheria  but  non-toxic 

I 


114  MICROBES   AND  TOXINS 

are  to  be  found ;  the  name  of  pseudo-diphtheria  has  been 
agreed  upon  for  them.  Are  they  capable  of  becoming  toxic  ? 
Under  the  conditions  in  which  we  can  experiment  it  is 
scarcely  possible  to  give  an  answer.  There  is  a  possibility, 
and  even  a  probability.  But  once  the  natural  selection  has 
occurred,  it  is  undoubtedly  the  toxic  bacilli  which  get  all  the 
chances  of  passing  from  mouth  to  mouth  and  of  maintaining 
their  hereditary  privileges. 

The  specificity  of  the  meningococcus  has  had  to  be  defended 
against  a  group  of  bacteria  resembling  it  in  form  and  cultural 
characters — the  pseudo-meningococti.  The  true  meningococcus 
may  be  distinguished  from  these  by  various  biological  reactions, 
but  it  is  certain  that  the  principal  character  from  our  point  of 
view  is  the  property  of  causing  meningitis,  and  it  is  difficult  to 
say  whether  or  not,  and  under  what  conditions,  the  "  pseudo  " 
forms  may  acquire  this  power. 

Not  only  have  there  been  distinguished  three  pathogenic 
types  of  the  dysentery  bacillus  (Shiga,  Flexner,  and  Strong), 
but  a  whole  group  of  pseudo- dysentery  bacilli  has  been 
admitted,  and  side  by  side  with  the  true  bacillary  dysentery 
there  have  been  decribed  dysenterifcrm  affections  caused  by 
these  "  pseudo-bacilli."  In  reality  the  biological  reactions 
have  proved  that  there  is  only  one  fundamental  type  of  patho- 
genic dysentery  bacillus,  but  that  there  exist  none  the  less 
satellites  of  this  which  possess  biological  importance,  although 
of  less  importance  from  the  point  of  view  of  medicine. 

Hygiene  cannot  afford  to  neglect  the  non-pathogenic 
cholera  vibrios,  often  very  difficult  to  distinguish  from  the  vibrios 
isolated  from  genuine  fatal  cases  of  cholera.  Since  the 
prophylaxis  is  based  on  the  discovery  of  the  microbe,  and 
since  no  laboratory  animal  exists  which  readily  takes  cholera, 
it  has  been  necessary  to  employ  refined  diagnostic  procedures, 
and  these  do  not  solve  the  scientific  question  of  the  relations 
in  nature  between  these  different  vibrios.  In  certain  maladies 
closely  resembling  typhoid  fever  (but  almost  always  benign, 
rarely  fatal),  bacilli  have  been  found  which  only  differ  from  the 
typhoid  bacillus  in  certain  cultural  peculiarities,  differing  also 


PATHOGENIC   MICROBES— INFECTION     115 

from  the  B.  coli  which  is  so  abundant  in  the  normal  intestine. 
Two  principal  types  can  be  distinguished,  A  and  B  :  the  latter 
is  nearer  to  the  B.  coli,  the  former  to  the  B.  typhosus.  They 
are  called  para-typhoid  or  para-coli  bacilli,  but  do  not  on  that 
account  call  in  question  the  specificity  of  the  Typhoid  bacillus  ; 
the  latter  rather  appears  as  a  chosen  specimen  out  of  a 
numerous  family  containing  other  well-defined  pathogenic 
bacteria,  among  which  are  not  only  these  para-A  and  B,  but 
various  bacilli  producing  meat-poisonings,  diarrhoea  among 
animals,  and  the  pneumo-enteritis  of  pigs.  This  family  is 
even  the  one  in  which  the  biological  reactions  have  been 
found  most  suitable  for  establishing  fairly  definite  degrees  of 
relationship.  We  class  them  by  their  relations  to  the  human 
species,  putting  at  the  head  of  the  column  the  typhoid 
bacillus.  It  is  evident  that  if  we  were  calves  or  pigs  our  point 
of  view  might  be  somewhat  modified. 

One  of  the  properties  characterising  the  tubercle  bacillus  of 
Koch  is  its  acid-fastness  (it  takes  on  stains  with  difficulty,  but 
once  stained  by  the  suitable  colour,  it  resists  the  decolorising 
action  of  acids).  There  exist  numerous  acid-fast  bacilli  and 
even  numerous  bacilli  capable  of  producing  tubercles  in  the 
tissues.  Between  the  human  and  bovine  bacilli  and  the 
bacilli  found  in  grass,  in  manure,  and  even  in  smegma,  there  is 
a  long  series  of  intermediate  types.  Are  we  to  believe  that 
among  the  acid-fast  and  para-tubercle  bacilli  the  ancestor  of 
the  tubercle  bacillus  of  men  and  the  ox  is  to  be  found  ?  It  is 
rather  a  philosophical  question  and  escapes  experimental 
examination.  But  the  general  truth  of  the  Darwinian  ideas 
compels  us  to  this  belief.  According  as  one  is  physician  or 
veterinary  surgeon,  according  as  one  is  engaged  in  diagnosis  or 
in  treatment,  according  as  one  is  accustomed  to  think  as  a 
naturalist  and  to  class  all  living  beings  in  groups,  so  one  tends 
to  insist  on  differences  on  the  one  hand,  on  resemblances  on 
the  other.  The  production  of  tuberculin  and  the  re-inoculation 
of  tuberculous  lesions  in  series  in  a  given  species  are  methods 
of  differentiation  which  do  not  invalidate  the  existence  of  a 
great  natural  family. 

I    2 


116  MICROBES  AND  TOXINS 

Small-pox  and  vaccination  are  without  doubt  of  common 
origin;  Jennerian  vaccination  is  based  as  much  on  their 
relationships  as  on  their  differences.!  In  the  same  way  the 
distinction  between  the  human  bacillus  and  the  bovine  bacillus 
is  not  inconsistent  with  a  common'  origin.  The  numerous 
experiments  which  have  been  made  to  vaccinate  cattle  with 
the  human  bacillus  prove  that'  it  is  a  question  of  two  adapta- 
tions from  the  same  type,  and  i  unfortunately  they  are  not 
sufficiently  differentiated  for  one  to  be  sure  that  the  bovine 
bacillus  is  incapable  of  producing  in  man  a  tuberculosis,  not 
merely  local  and  benign,  but  general  and  fatal. 

The  specificity  of  bacteria  is  only 'relative,  but  it  suffices  for 
the  carefully  performed  bacteriological  diagnosis  which  is  so 
useful  in  medicine;  Instead  of  the  solitary  types  which  once 
seemed  to  constitute  the  whole  species,  we  know  now  varieties 
and  families  of  which  one  member  only  may  be  the  constant 
cause  of  a  definite  disease ;  and  this  is  of  advantage  to  both 
medicine  and  hygiene.  In  theory  it  is  evident  that  the  only 
point  of  view  to  be  accepted  is  that  of  the  plasticity  of 
microbial  species,  i.e.,  the  Darwinian  theory. 

Virulence'.— Virulence  is  in  the  first  place  the  capacity  in 
a  microbe  to  settle  and  develop  in  the  bodies  of  animals; 
secondly,  its  capacity  of  secreting  its  toxic  substances.  Even 
in  the  strictly  toxic  diseases,  such  as  tetanus,  the  intoxication  is 
not  the  whole  malady ;  there  is  a  preliminary,  the  penetration 
of  the  microbe,  which  may  or  may  not  find  suitable  conditions 
for  its  growth. 

Virulence  is  a  variable  property,  and  it  was  in  connection 
with  modifications  in  virulence  that  Pasteur  had  the  intuition 
of  the  possibility  of  attenuating  a  virus. 

Diminution  of  Virulence.— In  cultures  on  artificial 
media  in  the  laboratory  the  virulence  'dimiflisn.es  spontaneously; 
the  media  may  be  improved  by  adding  ^animal  fluids  (serum, 
blood,  ascitic  fluid).  A  little  too  much  or  too  little  acidity  or 
alkalinity,  too  much  or  too  little  peptone  or  salt,  may  cause 
our  nutrient  broth  to  lower  the  virulence  of  the  strain  from 
the  change  in  its  reaction. 

The  diphtheria  bacillus  and  the  streptococcus  are  not  suited 


PATHOGENIC  MICROBES— INFECTION    1IT 

by  acids ;  the  cholera  vibrio  finally  suffers  from  the  alkalinity 
which  it  itself  produces;  the"  presence  of  fatty  materials  is 
injurious  to  anthrax  bacillus.  There  exist  processes  for 
diminishing  virulence :  i.  The  action  of  a  high  temperature 
(Toussaint,  Pasteur),  e.g.,  a  temperature  of  41°  to  43°C., 
instead  of  the  body  temperature  of  s6-37°C.  2.  Temperature 
+  aeration  (Pasteur's  experiments  on  fowl-cholera  and  swine- 
erysipelas).  3.  Desiccation  (Pasteur  :  preparation  of  the  spinal- 
cord  of  rabbits  in  the  treatment  of  hydrophobia).  4.  Light, 
pressure,  oxygen  under  pressure.  5.  Antiseptics  (Roux : 
carbolic  acid,  potassium  bichromate,  etc.). 

Increase  of  Virulence— Passages.— For  this  purpose 
one  provides  a  microbe  with  the  food-conditions  which  suit  it 
best;  oxygen  of  the  air  for  the  B.  diphtheria;  extracts  of 
putrefying  meat  for  the  B.  of  tetanus  (Brieger  and  Cohn). 
The  virulence  is  augmented  by  accustoming  the  bacteria  to 
the  body  against  which  they  are  being  prepared ;  the  feebler 
individuals  are  destroyed  by  the  natural  defences,  and  a 
selection  of  the  strongest  members  takes  place. 

A  bacterium  may  be  habituated  to  the  guinea-pig  by  com- 
pelling it  to  live  in  the  peritoneum  of  this  animal  enclosed  in 
a  collodion  sac,  which  permits  the  penetration  of  the  nutrient 
juices  while  keeping  off  the  leucocytes.  It  is  a  culture  in  the 
living  body.  Habituation  is  chiefly  produced  by  the  method  of 
passage,  i.e.,  by  inoculating  the  bacterium  in  an  animal  and 
from  this  animal  into  another  (generally  of  the  same  species). 
In  certain  cases  a  degree  of  virulence  is  reached  which  cannot 
be  exceeded  in  the  species  of  animal  employed  ;  thus  the  virus 
of  hydrophobia  becomes  virus  fixe  after  a  certain  number 
of  passages  through  the  rabbit.  Passages  do  not  perceptibly 
affect  the  tubercle  bacillus,  but  have  a  pronounced  influence 
on  the  streptococcus.  According  to  Marmorek  the  strepto- 
coccus, which  required  at  first  a  dose  of  i  c.c.  to  kill,  could  be 
brought  by  passage  to  kill  with  a  dose  of  o'ooooooooooi  c.c. 
Pasteur  raised  the  virulence  of  the  anthrax  bacillus  by  passing 
it  through  new-born  animals,  then  through  older  ones,  then 
adults,  and  finally  through  different  species. 

Passage  does  not  give  the  same  results  in  all  cases,  and 


118  MICROBES   AND   TOXINS 

qualitative  variations  occur  which  render  it  necessary  to  take 
into  account  the  species  of  animal  employed.  Pasteur,  after 
exalting  the  virulence  of  the  virus  of  rabies  for  the  dog  by 
passage  through  rabbits,  found  that  passage  through  monkeys 
weakened  it.  The  bacillus  of  swine-erysipelas  becomes  more 
virulent  (for  the  pig)  after  passage  through  pigeons,  less  virulent 
by  passage  through  rabbits.  Passage  through  a  foreign  species 
has  been  used  to  prepare  vaccines  ;  thus  the  virus  of  vaccine 
lymph — undoubtedly  of  the  same  origin  as  that  of  smallpox — 
has  become  a  "vaccine"  by  passage  through  the  cow;  the 
pox  of  pigeons  becomes  a  "  vaccine  "  for  pigeons  by  passage 
through  the  fowl.  In  passing  from  fowl  to  fowl  the  spirillum 
of  Marchoux  and  Salimbeni  gets  weaker.  And  while  it  is  true 
that  a  feebly  virulent  anthrax  strain  can  "  recover  "  its  virulence 
by  passages  beginning  on  new-born  mice,  in  the  spirillosis 
of  fowls,  on  the  contrary,  according  to  Marchoux,  it  is  precisely 
the  young  of  the  species  concerned  which  is  the  animal  of 
choice  for  weakening  the  virus  and  preparing  an  efficient 
vaccine  for  the  adult. 

Further,  the  age  of  the  "  young "  individual  must  be 
reckoned  precisely  :  it  is  known  that  the  new-born  child  is 
less  susceptible  to  Jennerian  vaccination  than  the  child  of 
three  or  four  months. 

The  modification  of  virulence  may  be  expressed  by  a 
difference  in  dose  in  relation  to  a  "  soil "  agreed  upon.  The 
example  of  Marmorek's  streptococcus  gives  a  numerical 
measure  of  the  increase  in  virulence.  The  spirochaete  of 
Schaudinn  was  inoculated  from  man  into  anthropoid  apes, 
from  these  into  the  lower  monkeys,  thence  into  the  rabbit, 
and  thence  into  the  guinea-pig ;  but  this  has  chiefly  been  a 
case  of  progress  in  the  technique  of  inoculation,  for  nowadays 
it  is  possible  to  inoculate  directly  from  man  into  the  rabbit. 

Attenuation  of  Viruses. — There  was  in  Pasteur's 
laboratory  a  culture  of  the  bacterium  of  fowl  cholera  which 
was  being  reinoculated  daily  and  was  of  constant  virulence. 
It  happened  that  a  culture  was  taken  for  inoculation  which 
had  remained  untouched  for  several  weeks  in  the  incubator ; 


PATHOGENIC   MICROBES— INFECTION     119 

the  fowls  became  ill  but  did  not  die,  and  further,  they  resisted 
a  second  inoculation  of  a  very  virulent  strain  which  killed  the 
controls.  This  was  the  first  demonstration  of  an  attenua- 
tion produced  by  keeping  in  contact  with  air  at  incubator 
temperature.  By  taking  cultures  of  different  ages,  a  scale  of 
virulence  could  be  produced— a  series  of  "vaccines." 

If  on  re-inoculating  an  enfeebled  culture,  one  obtained  a 
virulent  culture,  it  would  not  be  correct  to  speak  of  attenua- 
tion, but  merely  of  enfeeblement — transitory  lowering  of  the 
virulence. 

Attenuation  is  only  applied  to  a  permanent  enfeeblement, 
one  which  passes  from  one  generation  to  the  next.  One 
cannot  say  hereditary  because,  strictly  speaking,  heredity  only 
occurs  when  there  is  sexual  reproduction.  In  the  experiments 
on  fowl  cholera  the  new  cultures  showed  themselves  to  be 
weakened  in  series  ;  it  was  thus  a  true  attenuation. 

"  If  you  take  each  one  of  the  cultures  whose  virulence  has 
been  attenuated  as  the  starting  point  of  successive  cultures 
and  without  a  perceptible  interval  in  the  starting  of  the 
cultures,  the  whole  series  will  reproduce  the  attenuated 
virulence  of  the  one  serving  as  starting  point.  Similarly,  a 
culture  of  zero  virulence  reproduces  another  of  the  same" 
(Pasteur). 

A  similar  process  of  attenuation  appeared  at  first  inapplic- 
able to  the  anthrax  bacillus ;  as  the  culture  grows  older  the 
bacterium  sporulates  and  the  spores  are  not  affected  by  the 
conditions  which  act  upon  the  bacillary  form.  One  could  only 
therefore  expect  attenuation  from  an  anthrax  bacillus  which 
did  not  produce  spores. 

Now  at  42°'5  the  anthrax  bacillus  does  not  sporulate. 
Pasteur  cultivated  it  therefore  at  42°'$  in  order  to  diminish  its 
virulence  by  the  action  of  the  heat  and  the  air. 

It  was  then  found  that  sporulation,  instead  of  being  an 
obstacle  to  attenuation,  was  a  condition  entirely  favourable ; 
reinoculated  at  35°,  a  bacillus  attenuated  at  42° '5  produced 
sporulating  bacilli,  but  the  bacilli  germinating  from  these 
spores  possessed  the  same  degree  of  virulence  as  the  bacilli 


120  MICROBES   AND  TOXINS 

from  which  the  spore  was  derived.  Pasteur  had  obtained  an 
attenuation  fixed  by  the  resistant  form,  the  spore ;  "  vaccine 
viruses  fixed  in  their  germs,  with  all  their  peculiar  qualities 
without  any  possible  alteration." 

It  was  by  these  modifications  of  virulence  that  Pasteur 
explained  the  behaviour  of  the  great  epidemic  diseases  : 

"  There  exist  virulent  diseases  which  appear  spontaneously  in 
every  country:  such  is,  for  example,  the  typhus  fever  of  armies 
in  the  field.  Without  doubt  the  germs  of  the  microbes 
responsible  for  these  maladies  are  to  be  found  everywhere. 
Man  may  carry  them  on  his  body  or  in  his  alimentary  canal 
without  suffering  great  harm,  but  they  are,  nevertheless,  ready 
to  become  dangerous  when,  under  conditions  of  overcrowding 
and  successive  development  on  the  surface  of  wounds,  in 
weakened  bodies  or  otherwise,  their  virulence  becomes  progres- 
sively reinforced. 

"  Virulence  thus  appears  under  a  new  light  to  us  and  one 
which  is  distinctly  disquieting  for  mankind,  unless  nature  during 
the  past  centuries  has  already  met  with  all  the  opportunities 
possible  of  developing  virulent  or  contagious  diseases,  which 
is  highly  improbable. 

"A  microscopical  organism,  harmless  for  man  or  for  a  given 
animal  species,  is  simply  a  creature  which  cannot  develop  in 
our  bodies  or  in  the  body  of  the  given  animal,  but  there  is 
nothing  to  prove  that  if  this  microscopical  creature  succeeded 
in  penetrating  another  of  the  many  thousand  species  in  creation, 
it  might  not  invade  it  and  produce  in  it  disease.  Its  virulence, 
reinforced  then  by  successive  passages  through  individuals  of 
this  species,  might  become  powerful  enough  to  attack  such 
and  such  an  animal  of  higher  position,  man,  or  the  domestic 
animals.  In  this  way  it  is  possible  to  create  new  virulences 
and  new  contagions.  I  am  much  inclined  to  think  it  is 
thus  that  there  have  appeared  throughout  the  ages  small-pox, 
syphilis,  plague,  yellow  fever,  etc.  .  .  .  and  further  that  it  is 
by  phenomena  of  this  kind  that  there  from  time  to  time 
appear  certain  great  epidemics  ..." 


PATHOGENIC   MICROBES— INFECTION     121 


INFECTION 

Infection  may  be  defined  thus :  the  attack  on  one  living 
being  by  another  which  penetrates  it  and  lives  parasitically  at 
its  expense. 

It  is  simply  one  case  of  the  universal  struggle  and  com- 
petition among  the  species. 

The  conflict  between  the  invader  and  the  invaded  resolves 
itself  into  a  question  of  nourishment  and  digestion.  "The 
parasite  attacks  by  secreting  toxic  or  dissolving  substances, 
and  defends  itself  by  paralysing  the  digestive  and  expulsive 
powers  of  its  host.  This  latter  exerts  a  noxious  effect  on  its 
aggressor  by  digesting  it  or  eliminating  it  from  its  body,  and  it 
too  defends  itself  by  means  of  secretions." l 

Infection  exists  among  the  amoebae.  An  amoeba  invaded 
by  the  parasites  described  by  Metchnikoff  under  the  name  of 
Microsphczra  finally  succumbs.  Certain  infusoria  are  infected 
by  Acinetians,  which  pierce  their  cuticle  and  invade  them. 
The  green  euglena  is  subject  to  infection  by  lower  fungi  of 
the  chytridian  group ;  they  lose  their  green  chromatophores 
and  become  literally  anaemic.  Infectious  diseases  are  not  the 
peculiar  privilege  of  man  and  the  higher  vertebrates. 

On  the  great  problem  of  the  origin  of  microbes,  their  mode 
of  transmission,  and  the  way  in  which  they  penetrate  the  body, 
bacteriology  and  hygienic  science  have  accumulated  many 
facts. 

The  microbes  inhabit  the  air,  water,  the  soil,  animals,  and 
plants,  and  gain  access  to  the  patient  either  directly  by  simple 
contact  with  another  patient  or  thanks  to  more  or  less  numerous 
and  various  intermediate  agents.  Contact  is  sufficient  for  the 
transmission  of  measles,  small-pox,  and  scarlatina ;  these  are 
the  contagious  diseases  properly  speaking.  The  air  may  carry 
the  germ  from  one  individual  to  another.  In  the  air  the 
B.  tuberculosis  may  float,  attached  to  particles  of  dried  dust 
or  to  moist  droplets,  projected  into  the  air  by  the  patient 

1  Metcbnikoff,  Pathologic  comparte  de  F  Inflammation, 


122 


MICROBES   AND   TOXINS 


during  speech  or  coughing  (Fliigge).     Water-supplies  convey 
the  cholera  vibrio  and  the  typhoid  bacillus.    Along  with  garden 


natfetis 


Fig.  40. — Amoeba   Amoeba  viriais). 
(After  Gruber.) 


FIG.  41. — An  amoeba  dying  full 
of  parasites. 


SphcuwptuyQ 
applying  .its  fir 
loihe  infitsortan 


ct&tfidium 


FIG.  42. — Green  euglena  enclosing 
.     a    chytridium   (lower   fungus). 
(Metchnikoff.) 


^  two  parasitic 

' ~ 


FIG.  43. — Infusorian  attacked  by 
acinetian  parasites  (Spfuzro- 
phrya).  (Metchnikoff.) 


earth  the  tetanus  spore  may  gain  access  to  a  wound ;  the  soil 
may  spread  anthrax  spores  over  sheep  pastures.  The  foods 
which  are  eaten  raw  (milk,  meat,  vegetables)  convey  what  they 
have  gathered  from  the  soil  or  from  the  bodies  of  animals. 


PATHOGENIC   MICROBES— INFECTION     123 

The  vehicle  of  transmission  may  be  a  living  creature  instead  of 
an  inert  object ;  fleas  carry  plague  from  rat  to  rat  and  from 
man  to  man.  The  intermediate  inoculating  agent  may  con- 
stitute a  storehouse  and  even  a  culture  chamber  for  the  virus, 
e.g.,  the  tick  in  spirillum  fever.  This  intermediate  agent 
becomes  properly  speaking  a  host  when  the  germ  undergoes 
in  it,  and  can  only  undergo  in  it,  a  cycle  of  changes  by  which 
it  attains  the  stage  at  which  it  can  infect  us  ;  for  example,  the 
mosquito  for  the  parasite  of  malaria  and  the  tse-tse  fly  for  the 
trypanosome  of  sleeping-sickness.  The  transmission  of  bovine 
piroplasmosis  from  ox  to  ox  is  conducted  by  two  individuals : 
a  tick  becomes  infected,  produces  larvae,  and  these  larvae 
inoculate  another  ox. 

Latent  microbism  is  said  to  exist  when  the  organs  and  tissues 
contain  germs  which  remain  for  a  longer  or  shorter  time 
unsuspected. 

Pasteur  thought  that  our  organs  and  tissues  were  normally 
aseptic  :  "  The  human  body  is  completely  closed  to  the  intro- 
duction of  the  germs  of  fermentation  "  ;  and  he  was  still  more 
right  in  adding,  "  Except  the  alimentary  canal  and  again  except 
in  certain  pathological  conditions."  The  body  defends  itself 
well  against  the  microbes  which  enter  it ;  "  latent "  microbes 
can  only  be  those  which  escape  phagocytosis  at  least  temporarily. 
The  best  example  is  furnished  by  cases  of  spontaneous  tetanus, 
appearing  under  the  influence  of  a  heatstroke ;  this  tetanus  is 
due,  according  to  Vincent,  to  the  germination  of  spores  which 
have  penetrated  the  body  by  some  unknown  path,  and  have 
remained  there  for  several  days  or  even  several  weeks,  till  the 
day  when  the  excessive  heat  accompanied  by  fatigue  interrupted 
the  phagocytic  defence.1  The  experiments  of  Porcher  and 
Desoubry  teach  us  that  the  blood  is  rarely  aseptic  during 
digestion. 

Auto-infection  is  said  to  exist  when  an  individual  is  infected 

1  Tetanus  spores  inoculated  in  the  blood  or  under  the  skin  of  rabbits  are 
only  eliminated  after  three  or  four  weeks  (subcutaneous  inoculation),  or 
even  after  three  months  (intravenous  inoculation).  They  'wake  up'  and 
germinate,  should  favourable  conditions  supervene,  among  others  necrosis 
in  the  tissues  (Tarozzi). 


124  MICROBES   AND  TOXINS 

by  -bacteria  of  which  he  is  himself  a  carrier.  The  term  of 
atttO'infection  gained  a  precise  signification  only  when  germ- 
carriers  became  known. 

Germ-carriers  are  those  individuals  who  harbour  the  microbe 
of  the  disease  yet  present  no  symptom  of  this.  The  fact  is  not 
entirely  novel  (it  has  been  known  for  long  that  the  pneumo- 
coccus  exists  in  individuals  recovered  from  pneumonia,  and 
Pasteur  discovered  it  in  the  saliva  of  a  child  dead  of  rabies), 
but  it  is  only  during  recent  years  that  all  its  importance  has 
been  recognised. 

There  exist  carriers  of  typhoid  and  paratyphoid  bacilli,  of 
diphtheria,  of  cholera,  of  dysentery  and  of  meningococcus 
There  are  individuals  who  have  acquired  the  microbe  without 
yet  contracting  the  disease — precocious  carriers  :  others  recently 
cured  and  not  yet  freed  from  their  bacteria — convalescent 
carriers  ;  individuals  cured  weeks,  months,  or  even  years  before- 
chronic  carriers :  finally  there  are  occasionally  individuals  who 
have  never  had  the  disease — healthy  carriers  or  "  paradoxical 
carriers."  It  is  of  great  importance  to  recognise  germ  carriers 
especially  in  communities  where  the  general  life  is  intimate  and 
confined,  as  in  schools  and  barracks. 

In  typhoid  fever  in  particular  the  germ-carriers  are  most 
often  women.  According  to  Frosch's  statistics,  there  is  one 
woman  for  every  five  cases  of  typhoid,  but  of  every  five  chronic 
typhoid-carriers  four  are  women.  The  typhoid  bacilli  in 
chronic  typhoid-carriers  select  as  their  favourite  habitat  the 
bile-ducts. 

Conditions  of  Infection. — These  are  very  variable, 
varying  with  the  virulence,  the  resistance  of  the  body,  and  the 
method  of  inoculation. 

The  number  of  attacking  bacteria  can  be  calculated  fairly 
closely  in  experiments,  but  in  the  natural  disease  it  is  impossible 
to  tell  how  many  bacilli  are  necessary  to  determine  a  given 
infection. 

The  rabbit  is  extremely  sensitive  to  the  bacilli  of  fowl- 
cholera.  Now,  according  to  Watson  Cheyne,  if  10,000  to 
30,000  bacteria  are  injected  there  is  only  a  local  abcess ;  above 


PATHOGENIC   MICROBES— INFECTION     125 

this  figure  a  general  infection  is  practically  certain ;  but  on  the 
other  hand  the  same  observer  states  that  one  bacillus  is 
sufficient  to  produce  fatal  anthrax  in  the  mouse.  There  are 
certain  experimental  facts  as  regards  the  smallest  quantity  of 
bacilli  capable  of  producing  tuberculosis  but  these  have  not 
an  absolute  value.  H.  Buchner  found  that  to  spray  zooc.c.  of 
a  dilution  of  i  per  100,000  of  tuberculous  sputum  was 
sufficient  to  give  miliary  tubercle  of  the  lung  to  guinea-pigs. 
The  sputum  employed  contained  approximately  80,000  bacilli 
per  c.c.  and  he  calculated  that  each  guinea-pig  had  been  exposed 
to  the  attack  of  100 'bacilli  (these  figures  are  of  course  very 
approximate).  In  the  experiments  of  Preisz,  j-J^j  milligram  of 
sputum,  containing  about  forty  bacilli,  was  found  sufficient. 
In  the  experiments  of  Findel,  guinea-pigs  inhaling  about  sixty 
bacilli  regularly  took  tuberculosis.1 

Microbial  Associations. — The  microbes  which  cause 
diseases  are  never  derived  from  pure  cultures.  Their  virulence 
is  modified  not  only  by  the  chemical  properties  of  the 
medium  and  the  "  soil "  on  which  they  fall,  but  by  the 
presence  of  other  species  favourable  or  the  reverse.  There 
are  streptococci  which  aggravate  diphtheritic  sore  throats. 
The  coliform  bacillus,  the  torula,  and  the  sarcina  described  by 
Metchnikoff  favour  the  production  of  intestinal  cholera.  In 
tetanus  the  associated  bacteria  are  helped  also  by  other 
favouring  factors,  such  as  splinters,  bruising  of  the  tissues,  and 
blood  clots.  It  was  formerly  thought  that  the  streptococcus  of 
erysipelas  was  antagonistic  to  the  anthrax  bacillus.  Pasteur 
observed  an  antagonism  between  the  _B.  pyocyaneus  and  the 
anthrax  bacillus  ;  if  these  two  bacteria  are  inoculated  on  a 
medium  in  lines  which  cross  each  other,  the  anthrax  bacillus 
grows  very  feebly  at  the  points  of  intersection.  The  proteolytic 
enzyme  of  the  B.  pyocyaneus,  the  pyocyanase,  is  injurious  to 
many  bacteria,  and  occasionally  plays  the  part  of  a  disinfectant 
(Ernmerich). 

1  Tuberculous  sputum  (caseous  material)  frequently  contains  50,000 
bacilli  per  mg.  In  cultures  half-dried  on  paper  there  are  about  35  to  40* 
millions  per  mg. — with  a  possible  error  of  one  million  (Chausse).  /. 


126  MICROBES  AND  TOXINS 

In  the  infections  due  to  a  well  characterised  microbe, 
capable  of  itself  of  causing  the  disease,  the  associated  bacteria 
most  often  act  by  turning  upon  themselves  the  phagocytic 
attack  ;  their  role  is  thus  secondary. 

The  study  has  hardly  commenced  of  those  bacterial  associa- 
tions which  have  no  specific  pathogenic  power,  but  which  act 
nevertheless  favourably  or  the  reverse  by  the  products  of  their 
metabolism.  Their  sphere  is  chiefly  the  alimentary  canal,  and, 
according  as  the  dominant  flora  of  the  intestine  is  acid  pro- 
ducing or  produces  indols  and  phenols,  the  general  health 
escapes  or  is  subject  to  the  action  of  the  sclerosing  toxins. 
These  microbial  associations  thus  constitute  a  certain  condition 
or  disposition  rather  than  a  true  disease.  There  can  be 
distinguished  in  it  a  fundamental  flora  and  an  ace  idental  flora, 
and  these  can  be  modified  by  fortifying  one  species  at  the 
expense  of  the  others ;  it  is  in  this  that  bacteriotherapy 
consists. 

In  the  mouth,  microbial  associations  produce  a  disposition, 
more  or  less  marked,  towards  the  development  of  inflammations 
of  the  throat. 

The  body  presents  a  field  capable  of  infinite  variations ;  we 
have  to  reckon  with  species,  age,  and  physiological  conditions : 
hunger,  cold,  and  fatigue.  Experiment  alone  could  teach  us 
that  a  mammal,  such  as  the  rabbit,  is  more  sensitive  to  avian 
tuberculosis  than  to  the  tuberculosis  of  mammals,  or  that  the 
rabbit  is  extremely  sensitive  to  the  bacillus  of  fowl-cholera  and 
the  pigeon  to  that  of  swine-erysipelas.  The  Algerian  sheep  is 
more  resistant  to  sheep-pox  than  the  sheep  of  Camargue.  In 
general  very  young  animals  are  more  sensitive  than  adults, 
yet  the  young  pig  hardly  ever  contracts  swine-erysipelas  under 
three  months. 

Hunger,  heat,  or  cold,  and  fatigue  act  by  depressing  the 
phagocytic  defences. 

Paths  of  Penetration  into  the  Body. — The  mosquito 
inoculates  the  virus  which  it  carries  either  under  the  skin 
or  directly  into  a  blood-vessel.  The  spirochaete  produces 
syphilis  only  when  it  is  inoculated  strictly  in  the  subcutaneous 


PATHOGENIC   MICROBES— INFECTION     127 

cellular  tissue.  The  virus  of  hydrophobia  spreads  from  the 
region  of  the  bite  to  the  nerve  centres  along  the  nerve  trunks. 
Tuberculosis  appears  different  according  as  it  is  inoculated 
along  one  path  or  another ;  in  natural  disease  different  methods 
of  propagation  are  associated.  The  two  following  ideas  are  of 
great  importance. 

1.  The  role  of  the  intestine  in  those  diseases  in   which   the 
infection  is  not  purely  intestinal. — The  question  was  long  ago 
put  by  Chauveau   in   relation  to   tuberculosis.     Behring  has 
taken  it  up  again,  and  maintains  that  every  case  of  pulmonary 
tuberculosis  in  the  adult  is  the  extension  of  an  intestinal  tuber- 
culosis acquired  from  milk  in  the  earliest  infancy,  but  remain- 
ing latent  for  years.     Tuberculosis  in  general,  according  to 
him,  does  not  attack  the  lung  until  it  has  pierced  the  barrier 
of  the  intestine.     A  similar  origin  has  been  claimed  for  other 
infections  which  finally  settle  in  the  lung,  such  as  the  pneumo- 
coccal  inflammation.     A  comparative  case  was  found  in  the 
anthracosis  of  coal-miners  (the  impregnation  of  the  lung  with 
coal-dust),  and  since  it  is  easier  to  experiment  with  inert  dust 
particles  than  with  virulent  bacteria,  anthracosis  has  become 
the   field   of  study   and  discussion  in  the  question  for  and 
against  intestinal  infection. 

It  has  been  settled  by  experiment  that  living  bacteria  can 
pass  through  the  intestinal  mucosa  like  dust  particles  without 
leaving  any  lesion  as  a  mark  of  their  passage  But  such 
passage  only  occurs  as  a  rule  when  massive,  repeated  doses  are 
ingested,  and  when  there  are  in  the  intestine  such  injuries  as 
favour  penetration.  It  is  therefore  chiefly  by  inhalation  and 
by  penetration  of  the  lymphatics  and  blood-vessels  in  the 
neighbourhood  of  the  pharynx  that  the  tubercle  bacillus 
reaches  the  lungs. 

2.  Seats  of  election    and  receptive   cells. — The  hydrophobia 
virus  fixes  itself  on  the  nerve-tissue,  the  parasite  of  malaria  on 
the  red  corpuscle  of  the  blood,  other  protozoa  on  the  white 
corpuscles.      The   dysentery   bacillus,    inoculated    under    the 
skin,  proceeds  to  make  a  home  for  itself  in  the  large  intestine. 
The    bacillus    of   swine-erysipelas    inoculated   in   the   pigeon 


128  MICROBES   AND  TOXINS 

is  chiefly  found  infecting  the  large  endothelial  cells,  the 
"  Kuppfer  cells,"  of  the  capillaries  of  the  liver. 

If  the  virus  of  small-pox  or  of  sheep-pox  is  injected  intra- 
venously, it  is  simply  carried  by  the  blood  and  settles  and 
multiplies  in  the  skin.  In  those  diseases  which  Borrel  has 
grouped  together  under  the  name  of  Epithelioses,  there  exists 
a  fairly  strict  cellular  specificity :  the  virus  only  develops 
vigorously  in  the  interior  of  epidermal  cells,  and  these  cells 
from  the  moment  of  infection  take  on  a  special  character; 
they  are  therefore  justly  called  the  receptive  cells.  The  reason 
why  we  have  failed  to  inoculate  certain  diseases  is  that  the 
proper  site  of  inoculation  has  not  yet  been  discovered,  i.e.,  the 
receptive  cell,  or  that  this  receptive  cell  requires  to  undergo, 
before  becoming  truly  receptive,  certain  modifications  which 
are  still  unknown  and  which  we  cannot  reproduce. 

"  In  cancer  our  methods  of  inoculation  in  a  normal  indi- 
vidual fail  to  strike  the  receptive  cells  and  to  transform  the 
normal  into  cancer  cells  :  it  is  this  which  constitutes  the  whole 
etiological  problem  of  cancer  "  (Borrel). 

Between  the  date  of  penetration  of  the  virus  and  that  of  the 
appearance  of  the  disease  there  is  a  period  of  incubation,  during 
which  the  virus  propagates  itself,  multiplies  and  affects  the 
cells  on  which  depend  the  symptoms.  The  duration  of  the 
incubation  varies  primarily  with  the  virus,  secondarily  with  its 
quantity  and  the  path  by  which  it  has  gained  access.  For 
example,  the  incubation  is  quite  short  when  experimental 
septicaemia  is  produced  with  a  virulent  streptococcus,  whereas 
in  human  leprosy  it  may  last  for  years.  In  rabies  the  period 
of  incubation  is  shorter,  and  the  disease  more  violent  when 
the  bite  is  on  the  face  than  when  it  is  on  the  leg.  In  tetanus 
(a  toxin  disease),  the  longer  the  incubation  the  less  serious 
the  disease. 

The  spirillum  of  recurrent  fever  causes  a  disease  of  the 
septicaemic  type,  i.e.,  the  microbe  inhabits  the  blood.  The 
tubercles  characteristic  of  tuberculosis  and  glanders  represent  a 
reaction  of  the  cells  of  the  mesoderm,  while  the  pustular 
diseases  like  small-pox  are  typically  reactions  of  ectodermic 


PATHOGENIC   MICROBES— INFECTION     129 

cells.  But  the  same  disease  may  be  in  different  phases 
septicaemia  or  localised  in  a  tissue,  and  in  any  case  the  blood 
itself  is  simply  a  tissue  whose  cells  are  motile.  It  was  long 
thought  that  typhoid  fever  was  an  infection  localized  to  the 
small  intestine  and  to  the  Peyer's  patches  there,  but  in  reality 
it  is  septicasmic  during  the  whole  of  the  febrile  period.  There 
are  also  septicaemic  phases  in  pneumonia  and  tuberculosis. 

Microbes  are  not  inert  particles  but  living  cells  acting 
through  their  secretions  and  toxins.  Cholera  is  an  intestinal 
infection,  but  it  is  fatal  to  its  host  from  a  general  intoxication. 
Every  infection  is  to  some  extent  an  intoxication.  Even  the 
macroscopic  parasites,  to  which  formerly  only  a  physical 
activity  was  ascribed,  the  bothriocephalus,  the  ankylostoma, 
the  trichina,  secrete  poisons  which  have  been  studied. 

Microbes,  ferments,  and  toxins  are  inseparable  terms,  and 
for  this  reason  the  discovery  of  the  diphtheria  toxin  by  Roux 
and  Yersin  in  1888  began  a  new  era  in  bacteriology. 


CHAPTER  VI 

INFLAMMATION  AND  PHAGOCYTOSIS 

The  comparative  pathology  point  of  view — Inflammation  throughout  the 
series  of  natural  species — defined  by  phagocytosis — Inflammation 
among  invertebrates  without  nerves  or  blood-vessels — The  phagocytes 
and  intracellular  digestion  —  Chemiotaxis  —  Phagocytes  in  man — 
Phagocytosis  in  the  chronic  infections — The  examples  of  the  squirrel- 
rat,  spermophilus,  and  of  the  jerboa. 

THE  essential  fact  of  inflammation  is  the  reaction  of  the 
phagocytes  towards  the  injurious  material  (Metchnikoff). 
Phagocytosis  is  not  a  theory  but  a  doctrine,  a  collection  of 
accumulated  facts.  It  has  put  the  finishing  touch  to  the  work 
accomplished  in  medicine  by  Darwin,  Virchow  and  Pasteur. 
From  Darwin  it  has  derived  its  fundamentally  evolutional 
character  :  it  is  founded  primarily  on  comparative  pathology  and 
demonstrates  the  persistence  of  the  same  phenomenon 
throughout  all  the  animal  species.  It  has  derived  from  Virchow 
its  foundation  on  cellular  pathology,  i.e.,  on  the  essential  role  of 
the  body  cells  in  disease.  From  Pasteur  it  derives  the 
fundamental  idea  of  the  role  of  microbes  in  the  production  of 
infections. 

It  required  a  zoologist  applying  the  method  of  comparative 
study  to  demonstrate  that  the  only  constant  phenomenon  in 
the  different  forms  of  inflammation  is  the  active  incorporation 
of  injurious  elements  by  fixed  or,  more  often,  migratory  cells 
which  are  capable  of  digesting  these.  Inflammation  is 
essentially  phagocytosis  and  phagocytosis  is  summed  up  by 
intracellular  digestion. 

130 


INFLAMMATION  AND  PHAGOCYTOSIS     131 

The  four  cardinal  symptoms  redness,  heat,  pain  and  swelling 
represent  only  an  external  definition  of  inflammation.  The 
sum  total  of  the  facts  is  so  complex  that  many  observers  in 
former  days  refused  to  give  a  simple  definition,  proposed  to 
abandon  the  vague  term  of  inflammation,  and  limited  themselves 
to  a  description  of  the  variety  of  facts. 

The  tissues,  the  vessels,  and  the  nerves  of  the  injured  part 
participate  in  the  state  of  inflammation :  to  which  is  the 
primary  role  to  be  ascribed  ?  Virchow  maintained  that  it  was 
the  tissues,  and  that  these  were  in  a  state  of  supernutrition  at 
the  expense  of  \hzfluid  parts  of  the  blood ;  the  cells  multiply 
at  the  injured  point  and  it  is  from  the  tissues  of  this  same 
region  that  the  numerous  cells  of  the  inflammatory  exudate 
are  derived.  Inflammation  taken  as  a  whole  represents  a 
danger  to  the  body. 

But  when  Cohnheim,  in  his  observations  on  the  frog's 
mesentery  exposed  to  the  air,  discovered  diapedesis  or  the 
escape  of  the  white  corpuscles  through  the  walls  of  the 
vessels,  and  when  it  was  established  that  the  pus  cells  instead 
of  developing  on  the  spot  by  the  proliferation  of  the  cells  of 
the  connective-tissue  came  from  the  motile  cells  of  the  blood, 
the  primary  fact  of  inflammation  seemed  to  be  the  vascular 
irritation,  the  other  appearances  being  secondary.  Cohnheim 
thought  he  had  proved  this  by  his  well-known  experiment ;  a 
frog's  tongue  is  ligatured  at  the  base  so  as  to  stop  the 
circulation.  On  untying  at  the  end  of  forty-eight  hours  the 
circulation  is  re-established,  but  is  now  of  the  inflammatory 
type  with  diapedesis. 

But  if  the  vascular  phenomena  are  of  primary  importance 
it  is  difficult  to  account  for  the  fact  that  microbial  or  other 
foreign  substances  introduced  under  the  skin  produce  an 
inflammatory  reaction,  but  fail  to  do  so  when  injected  into  the 
vessels  themselves. 

Light  was  first  thrown  on  the  problem  by  the  comparative 
study  of  lower  organisms. 

Inflammation  in  the  Lower  Organisms. — The  jelly- 
like  plasmodium  of  a  mycetozoon  pricked  or  burnt  responds 

K    2 


132 


MICROBES   AND   TOXINS 


to  the  injury  by  movements  of  attraction  or  repulsion 
(varying  according  to  circumstances)  of  its  protoplasm.  A 
foreign  body  introduced  into  it  is  engulfed  and  then  rejected. 


FIG.  44. — Plant  filament  surrounded  by  the  phagocytes  of  Spongilla. 
(Metchnikoff.) 

If  into  the  body  of  a  sponge  a  little  glass  tube  or  an  asbestos 
fibre  or  any  sharp  foreign  substance  is  introduced,  motile 
amoeboid  cells  from  the  mesoderm  soon  come  and  surround  it, 
the  same  cells  which  are  capable  of  surrounding  and  digesting 
both  inert  granules  and  living  prey.  This  engulfing  power  of 

phagocytes 


FIG.  45. — Mass  of  phagocytes 
round  a  spike  in  Bipinnaria 
asterigeria.  (Metchnikoff.) 


FlG.  46.— Phagocytes  of  the 
worm  collected  round 
a  foreign  body.  (Metch- 
nikoff.) 


the  mesodermic  cells  (and  of  certain  endodermic  cells  also)  is 
aided  by  the  sensitive  contractile  ectodermic  elements. 

The  larvae  of  a  certain  sea-anemone  (Astroptcttn ptntacanthus\ 


INFLAMMATION  AND  PHAGOCYTOSIS     133 

transparent  and  thus  easily  observed,  have  neither  nervous 
system  nor  vessels  nor  muscles :  they  react  towards  a  pene- 
trating foreign  body  by  an  accumulation  of  amoeboid  meso- 
dermic  cells.  In  a  larger  larva,  Bipinnaria  asterigeria,  the  cells 
of  the  mesoderm  can  be  seen  engulfing  particles  of  carmine  or 
of  indigo  and  surrounding  a  splinter  or  a  drop  of  blood  with 
masses  of  cells  equivalent  to  a  plasmodium.  They  also  engulf 
bacteria  introduced  under  their  outer  skin. 

In   all   these  cases  inflammation  occurs  without  blood  or 
blood  vessels.     Among  the  Annelids  which  possess  a  closed 


monospotne 


FIG.  47. — Daphnia  infected  by  Monospora.    FIG.    48. — Different   stages  of 
(Metchnikoff.)  Monospora.     (Metchnikoff.) 

vascular  system,  the  reaction  against  foreign  bodies  takes  place 
in  the  same  way  without  any  intervention  of  the  blood  vessels. 

In  LumbricuS)  whose  male  sex-glands  are  infected  by 
gregarines,  there  is  a  struggle  between  the  two  organisms,  the 
gregarine  encysting  itself  and  becoming  surrounded  by  an  outer 
covering  of  protective  chitin,  while  the  amoeboid  cells  sur- 
rounding it  join  together  and  form  a  sort  of  armour-plating 
which  stifles  it.  The  blood-vessels  remain  inactive. 

For  an  example  of  parasitism  exactly  parallel  to  an  infectious 
disease  there  is  nothing  better  than  to  observe  Daphnia  magna 
being  invaded  by  a  microscopic  fungus,  the  Monospora  bicuspi- 
data.  The  motile  cells  engulf  the  spores  of  the  mould  and 


134 


MICROBES   AND  TOXINS 


attempt  to  destroy  them  by  digestion :  a  struggle  takes  place, 
sometimes  the  Daphnia  is  victorious,  sometimes  it  succumbs. 

In  the  young  of  a  vertebrate,  the  axolotl,  if  the  non-vascular 
rudiment  of  the  fin  is  pricked  with  a  needle  charged  with  a 
little  carmine  or  indigo  powder,  the  migratory  cells  can  be  seen 


FIG.  50. — Two  leucocytes  of  Daphnia  sur- 
rounding a  conidium  of  Monospora. 
(Metchnikoff.) 


filaments 
cfosc&aria 


FIG.  49. — Spores  of  Mono- 
spora, surrounded  by 
leucocytes  of  Daphnia. 
(Metchnikoff.)  The  spore 
is  transformed  into 
granules. 

FlG.  51. — An  amoeba  (Amoeba  verrucosa) 
incorporating  a  filament  of  Oscillaria. 
(After  Rhumbler.) 

hastening  to  the  injured  point  and  engulfing  the  particles.  In 
the  older  stages  of  the  axolotl  and  in  the  tail  of  tadpoles,  where 
a  well-developed  vascular  system  exists,  inflammation  is  accom- 
panied by  a  dilatation  of  the  vessels  and  by  diapedesis  :  the 
reaction  is  more  violent,  but  the  essential  process  is  the  same 
as  in  the  non- vascular  invertebrates. 

"  It  is  quite  evident  that  inflammation  in  the  vertebrate,  in 
which  the  protective  phagocytes  emerge  from  the  vascular 
system  to  attack  the  aggressor,  differs  from  the  analogous 
phenomena  in  the  invertebrate  only  from  the  purely  quantitative 
standpoint. .  .  .  The  morbid  phenomena,  properly  speaking,  such 
as  the  lesion  or  the  primary  necrosis,  equally  with  the  processes 
of  repair  which  succeed  the  inflammation,  do  not  belong  to  it 
and  must  not  be  confused  with  it "  (Metchnikoff). 

The  phenomena  of  vascular  dilatation  and  hyperaemia  are  no 


INFLAMMATION  AND  PHAGOCYTOSIS     135 

more  to  be  regarded  as  inflammation,  says  Cantacuzene  with 
justice,  than  the  congestive  phenomena  which  accompany 
ovulation  or  precede  coitus  can  be  called  fertilisation. 

Phagocytes,  phagocytosis  and  digestion. — Metchnikoff 
has  given  the  name  of  phagocytes  to  those  cells  which  are 
capable  from  their  own  activity  of  seizing  and  incorporating 
solid  particles.  (There  is  no  question  here  of  the  faculty  of 
absorbing  substances  in  solution.) 

Certain  phagocytes  are  migratory  cells  :  for  example  the  white 
corpuscles  of  the  blood.  Others  are  fixed  cells — for  example 
many  of  the  endothelial  cells  of  the  blood-vessels  and  lymphatics, 
the  endothelial  cells  of  the  omentum  and  the  neuroglia 
cells.  Others  originally  motile  may  become  fixed  at  a  certain 
period  in  their  existence. 

The  fundamental  property    of  phagocytes    is    intracellular 
digestion. 

Metchnikoffs  first  observations  were  on  the  property  of 
intracellular  digestion  in  the  intestinal  epithelium  of  a  great 
many  Turbellarians.  In  the  Ccelenterata  and  the  Sponges 
digestion  is  intracellular,  /.*.,  the  nutritive  particles  instead  of 
being  digested  in  a  cavity  with  the  help  of  juices  poured  out 
by  digestive  cells  undergo  digestion  in  the  interior  of  the  cells 
themselves. 

The  Metazoa  have  inherited  this  property  from  the  Protozoa. 
Originally  all  the  cells  of  the  inferior  Metazoa  are  capable  of 
phagocyting  ;  there  are  both  ectodermic  and  endodermic  pha- 
gocytes ;  later  the  function  devolves  entirely  on  specialized 
cells  belonging  to  the  mesoderm. 

The  phagocytic  cells  of  man  are  descendants  of  cells  whose 
normal  function  was  to  digest  intracellularly,  and  through  them 
we  still  possess  this  method  of  digestion,  side  by  side  with  the 
extracellular  digestion  which  occurs  in  our  stomach  and 
intestine. 

Phagocytosis  is  thus  a  function  very  wide-spread  among 
living  beings,  and  the  struggle  against  infection  is  only  a 
particular  case  of  it. 

"  The  phagocytes  are  those  cells  which  have  best  preserved 


136  MICROBES  AND   TOXINS 

the  primitive  amoeboid  type.  They  are  in  general  the  least 
differentiated  elements  in  the  body,  but  they  are  also  the  most 
independent  and  possess  the  greatest  vitality.  They  assist  in 
building  up  the  young  animal  during  the  embryonic  period,  and 
when  the  tissues  begin  to  wear  out,  when  old  age  is  coming  on, 
it  is  the  phagocytes  which  consume  the  senile  cells  incapable 
of  recovery  and  take  their  place.  The  renewal  of  cells  and 
tissues  which  goes  on  slowly  and  continuously  in  a  great  many 
animals  is,  like  the  abrupt  transformation  which  occurs  in 
metamorphosis,  the  work  of  phagocytes." 

The  importance  of  pathological  phagocytosis  from  the  medi- 
cal point  of  view  should  not  make  us  forget  that  a  normal 
phagocytosis  exists.  The  histolysis  in  the  larvae  of  insects,  the 
destruction  of  the  tail  in  the  tadpole  forms  of  the  Tunicata,  the 
degeneration  of  the  tail  muscles  in  the  tadpole  of  the  Frog  and 
Toad,  the  destruction  of  the  myelinated  nerve  fibres  in  Wallerian 
degeneration,  the  shrinking  of  the  ovarian  follicles,  the  fixation 
of  the  ovum  on  the  mucous  membrane  of  the  uterus,  the  daily 
destruction  of  the  red  corpuscles  of  the  blood  which  goes  on 
in  the  spleen,  all  are  examples  of  normal  phagocytosis. 

The  phagocytes  are  guided  or  directed  in  their  choice  and 
perception  of  the  bodies  which  they  ingest,  by  a  peculiar  sense 
whose  manifestations  are  known  by  the  name  of  chemiotaxis. 

It  has  been  known  since  the  time  of  Pfeffer  and  Stahl  that 
cellular  organisms  and  plasmodia  are  attracted  by  certain 
substances  (positive  chemiotaxis),  and  repelled  by  others 
(negative  chemiotaxis).  They  become  accustomed  to  sub- 
stances which  at  first  repelled  them  and  finally  are  attracted  by 
these.  Massart  and  Ch.  Bordet  have  systematically  studied 
chemiotactic  actions,  by  introducing  under  the  skin  of  the  frog 
capillary  tubes  containing  chemical  substances,  microbes  and 
their  products.  Lactic  acid,  glycerine,  bile,  and  guanin  repel 
the  leucocytes ;  sterilised  cultures  of  both  saprophytic  and 
pathogenic  microbes  attract  them.  Positive  chemiotaxis  may 
be  considered  as  the  appetite,  which  prepares  for  intracellular 
digestion.  Everything  is  not  yet  explained  in  this  distant 
action.  Chemiotaxis  is  analogous  to  the  sensations  of  higher 


INFLAMMATION  AND  PHAGOCYTOSIS     137 

animals,  and  the  sensations  of  a  plasmodium  obey  like  ours 
the  law  of  Weber. 

Chemiotaxis  is  a  sort  of  chemical  sense. 

The  phagocytes  have  also  a  sort  of  tactile  sensibility.  The 
leucocytes  in  their  reaction  apply  themselves  to  the  exciting 
body  over  the  largest  surface  possible. 

In  defensive  phagocytosis,  the  struggle  of  the  body  against 
the  parasitic  invaders,  it  is  not  necessary  to  suppose  any 
purposeful  cause  but  simply  a  function  developed  by  evolution 
and  selection.  "  Those  lower  animals  in  which  the  motile  cells 
directed  themselves  towards  the  enemy,  engulfing  and  destroying 
them,  survived,  whereas  others  in  which  the  phagocytes  did  not 
act,  were  condemned  to  perish.  All  the  useful  characters  and 
among  them  those  which  are  concerned  in  the  inflammatory 
reaction  have  become  fixed  and  transmitted  without  the  inter- 
vention of  any  preconceived  purpose  whatever"  (MetchnikorT). 
Thus  in  the  invertebrates  with  soft  skins  in  which  bacterial 
invasion  occurs  easily  there  has  been  a  selective  process  at 
work  in  the  phagocytic  apparatus  and  the  defensive  measures 
have  become  perfected.  Among  the  invertebrates  possessing 
a  natural  protection,  such  as  a  chitinous  covering,  infection  is 
rarer,  but  the  means  of  defence  have  not  found  suitable 
conditions  for  their  employment  and  development,  so  that  the 
infected  organism  succumbs.  The  phagocytic  arrangements 
are  much  reduced  in  the  Insects,  and  in  these  the  parasitic 
fungi  have  great  difficulty  in  penetrating  the  cuticle,  but  if 
they  are  successful  the  insect  is  destroyed  (for  example  in  the 
beetle  Cleonus  punctiventris  invaded  by  Isaria  destructor}.  The 
nematode  worms  which  are  protected  by  a  thick  skin  do  not 
even  possess  cells  capable  of  movement. 

The  phagocytes  of  man  are  both  fixed  and  motile.  Among 
the  fixed  are  the  large  mononuclears,  the  Kuppfer  cells  of  the 
liver,  certain  endothelial  cells  of  the  lung  (the  dust-cells)  and 
the  myeloplaxes  of  the  bone-marrow.  The  motile  phagocytes  are 
the  white  corpuscles  or  leucocytes  in  general  (except  the  small 
lymphocytes),  the  polymorphs,  the  eosinophils,  the  large 
mononuclear  cells  of  the  blood  and  of  the  lymphatic  organs. 


138 


MICROBES   AND  TOXINS 


Metchnikoff  has  divided  the  phagocytes  into  macrophages 
and  microphages ;  the  former  are  chiefly  concerned  in  the 
absorption  of  cells  and  cellular  debris,  and  include  the  large 
mononuclears,  the  fixed  phagocytes  of  the  spleen,  of  the 
peritoneum  and  of  the  lymphatic  glands.  They  digest  the 
blood  corpuscles  and  other  phagocytes.  The  microphages 

are  the  polymorphs ;  their 
principal  function  is  to 
digest  bacteria.  There  are 
exceptions.  In  certain 
cases  the  microphages  take 
up  cells  (red  cells  among 
others),  while  in  certain 
cases  the  macrophages 
take  up  bacteria ;  the  large 

FIG.    52.  -  Different  leucocytes.  - 1.      mononuclears  surround  the 
Polymorph.— 2.  Microphage  (poly-       tubercle  bacillus  producing 
morph)  taking  up  staphylococci,  tf.       th       •      t        n         d  tak 
—3.    Small  lymphocyte.— 4.    Eo- 
sinophil. — 5.    Large   mononuclear 
(macrophage).  —  6.     Macrophage 
from  the  peritoneum  of  a  guinea- 


pig  taking  up  red  corpuscles.  —  7. 
Macrophage  from  the  peritoneum 
taking  up  polymorphs  (micro- 
phages) and  blood  corpuscles. 


also  the  spirochaetes  of 
recurrent  fever  and  of 
syphilis. 

It  has  sometimes  been 
maintained  that  the  pha- 
gocytes only  take  up  dead 
bacteria  and  not  living  virulent  ones ;  this  is  a  mistake  to 
which  it  will  be  necessary  to  refer  again  in  connection  with 
immunity.  Under  the  microscope  there  can  be  seen  inside 
the  phagocytes  living  and  even  motile  bacilli,  and  cultures 
can  be  obtained  by  inoculating  into  broth  phagocytes  full  of 
microbes :  the  leucocytes  are  destroyed  and  the  liberated 
bacteria  multiply.  They  were  still  quite  alive  therefore, 
although  already  seized  by  the  phagocytes. 

The  phagocytes  secrete  digestive  ferments.  Rossbach  has 
demonstrated  the  existence  of  a  starch-splitting  ferment  in  the 
leucocytes  of  the  tonsils.  The  cells  of  pus  can  digest  fibrin 
and  gelatine,  and  must  thus  secrete  proteolytic  ferments.  In 
cases  of  acute  muscular  atrophy  the  progressive  digestion  of 


INFLAMMATION  AND  PHAGOCYTOSIS     139 

the  muscle  fibres  can  be  observed  in  the  interior  of  the 
phagocytes.  The  bacilli  or  cells  taken  up  by  the  phagocytes 
become  distorted,  and  before  disappearing  lose  their  affinity 
for  stains.  In  the  various  cases  of  immunity  the  phagocytes 
digest  the  bacteria  by  means  of  endo-enzymes. 

The  surface  of  the  skin,  and  in  particular  of  the  mucous 
membrane,  is  being  continually  besieged  by  bacteria,  and 
never  a  moment  passes  but  some  point  in  the  body  is  in  a  state 
of  subinflammation.  The  phagocytes  are  in  continual  opera- 
tion on  the  surface  of  the  tonsils,  of  the  mucous  membrane  of 
the  intestine,  and  of  the  alveoli  of  the  lungs. 

Phagocytosis  plays  a  pre-eminent  part  in  chronic  infections, 
especially  in  tubercle,  and  the  tubercle  itself  is  a  phagocytic 
formation.  In  contrast  to  Baumgarten's  contention  that  the 
tubercle  is  built  up  by  epithelial  cells,  fixed  cells  from  the 
diseased  tissue  itself,  lung,  liver,  or  kidney,  Metchnikoff 
and  his  pupils  have  proved  that  it  is  really  composed  of 
migratory  mesodermic  cells  which  have  come  from  elsewhere 
to  the  infected  point.  Borrel  followed  the  formation  of  the 
tubercle  from  the  time  of  the  first  contact  between  the  white 
corpuscles  and  the  bacilli,  and  found  that  injected  bacilli  were 
engulfed  by  the  polymorphs  while  still  in  the  circulation.  The 
polymorphs  perish  and  degenerate  (in  two  or  three  days)  and  are 
followed  by  the  macrophages  which  fuse  together  into  a  sort  of 
little  plasmodium  with  several  nuclei,  which  is  characteristic  of 
the  tuberculous  lesion,  and  is  called  by  the  anatomists  the 
giant  cell.  Later,  the  tubercle  may  soften,  and  there  may  be  a 
new  afflux  of  polymorphs  attracted  chiefly  by  the  bacteria  of  a 
secondary  infection. 

In  the  squirrel-rat  spermophilus,  a  rodent  rather  resistant  to 
tubercle,  the  phagocyted  bacilli  lose  their  staining  properties, 
degenerate,  swell  up  and  finally  appear  as  yellowish  bodies, 
such  as  are  never  observed  either  in  cultures  or  outside  the  cells, 
and  can  only  be  residues  of  phagocytic  digestion.  In 
another  rodent,  the  jerboa,  there  is  found,  especially  in 
tubercle  of  the  spleen,  instead  of  bacilli  amorphous  bodies 
built  up  of  concentric  layers,  which  are  encrusted  with 


140  MICROBES   AND   TOXINS 

phosphate  of  lime  and  may  be  dissolved  by  an  acid.  Obser- 
vation of  these  tubercles  at  different  stages  shows  that  the 
concentric  layers  correspond  to  secretions  of  the  bacillus  which 
has  been  defending  itself  against  the  phagocytes.  Analogous 
formations  are  known  in  actinomycosis  (the  club-forms  of 
the  granules).  There  is  no  essential  difference  between  the 
struggle  of  the  tubercle  bacillus  against  the  giant  cell  and  the 

Tubercle 
bacillus 


FIG.  54. — Giant  cell  enclos- 
pIG.    53.— Giant    cell    from    the  ing  the  final  stage  of  a 

spleen  of  the  jerboa  :   it  con-  calcareous  particle, 

tains  a  tubercle  bacillus  sur- 
rounded by  concentric  layers. 
(Metchnikoff.) 

struggle  of  the  gregarines  and  the  nematodes  (larvse  of  Gordius 
or  Rhabditis)  against  the  phagocytes  of  the  worm. 

Inflammation  is  thus  defined  by  phagocytosis  :  the  vessels 
and  nerves  have  their  importance,  but  are  merely  accessory. 
Infection,  inflammation,  and  immunity  can  all  be  seen  in 
miniature  in  the  examples  of  the  Bipinnaria  with  its  splinter 
surrounded  by  motile  cells,  and  of  the  Daphnia  with  its 
globules  in  the  act  of  devouring  the  spores  of  Monospora. 


CHAPTER  VII 

THE   PATHOGENIC  PROTOZOA  !    FILTER-PASSING  VIRUSES 

Protozoal  diseases— Laveran's  discovery— Importance  of  the  morphology 
and  the  life-cycle — Intracellular  protozoa — Heredity  in  bacterial  and 
in  protozoal  diseases — Diseases  due  to  the  so-called  invisible  microbes. 
— The  ultramicroscope — Filtration — Various  types  of  virus  capable 
of  passing  filters— Microbes  of  extreme  minuteness  described  in  the 
pustular  diseases  of  the  epithelium — Lesions  of  the  infected  cells. 

THE  PATHOGENIC  PROTOZOA 

THE  name  of  Pasteur  must  be  inscribed  at  the  head  of  this 
chapter.  It  was  by  his  study  of  pebrine,  a  protozoal  disease 
of  silk-worms,  that  our  ideas  on  the  microbial  diseases  were  so 
much  advanced. 

The  studies  on  anthrax,  the  labours  of  Koch  and  the  great 
discovery  of  the  attenuation  of  viruses  led  the  new  science  in 
the  direction  rather  of  bacteriology;  the  protozoa  had  even 
been  somewhat  forgotten,  when  in  1880  Laveran  discovered 
among  them  the  cause  of  malaria.  Since  that  date  their 
importance  in  pathology  has  never  ceased  to  grow. 

The  methods  of  research  cannot  be  quite  the  same  as  in 
bacteriology ;  they  have  not  the  same  simplicity  as  the  bacteria. 
In  the  case  of  the  tubercle  bacillus,  the  cholera  vibrio,  and  the 
streptococcus,  we  practically  know  only  one  single  constant 
fixed  form  for  each,  and  there  is  no  sign  of  a  life-cycle.  The 
majority  of  the  pathogenic  protozoa  on  the  contrary  go  through 
a  cycle  in  their  existence  whose  successive  forms  may  be  very 
diverse  and  this  cycle  may  take  place,  not  in  a  single  host,  but 
often  in  two  different  ones.  The  discoveries  of  Ross  on  the 

141 


MICROBES  AND  TOXINS 

plasmodium  of  malaria  present  the  best  example  of  the  labour 
necessary  to  reconstruct  the  biology  of  a  parasite  common  to 
man  and  the  mosquito. 

PROTOZOAL  DISEASES 
Rhizopods 

Amoebae. — Amoebic    dysentery  and  liver  abscess.     A  ciliated  infusorian, 
Balantidium  coli,  may  produce  the  same  disorders. 

Haematozoa 1 

Trypanosomes. — Sleeping  sickness  (human  Trypanosomiasis). — Trypano- 

somiasis  of  animals  :  nagana,  surra,  dourine,  mal-de-Caderas. 
Leishmania. — "  Leishmanioses "  :    Kala-azar    (the   oriental  sore,    Biskra 

button,  Aleppo  button,  etc.,  being  particular  cases). 
Piroplasmoses. — Bovine   Piroplasmoses   (due   to    Piroplasma   bigeminum, 

P.  parvum,  P.  mutans  ;  canine,  ovine,  and  equine  Piroplasmoses. 
Plasmodia. — Malaria  with   its  varieties:    tertian,    quartan,    and    sestivo- 

autumnal  or  tropical  tertian. 

Spirochsetes. — Relapsing  fever  (European,  African,  Asiatic,  American). 
The  spirillosis  of  fowls  and  geese. 
Human  spirillosis  =  syphilis. 

The  more  the  study  of  these  cycles  was  advanced,  the  more 
one  was  compelled  to  acknowledge  relationships  between  forms 

which  did  not  seem  in 
the  least  related.  Es- 
tablished classifications 
have  several  times  got 
into  difficulties  from 
trying  to  express  genea- 
logical relationships 
between  very  diverse 
forms.  Schaudinn  saw  in 
the  life-cycle  of  the  same 
parasite  trypanosomes, 
spirochaetes,  and 
amoeboid  forms.  It  has 
been  necessary  to  re- 
cognise a  relationship  between  the  sporozoa  (haemo-sporidia) 
and  the  flagellates.  The  affinities  of  the  spirilla  and  spiro- 

1  According  to  the  recent  views  of  Hartmann,  originating  in  Schaudinn's 
ideas,  all  the  haematozoa  mentioned  here  may  properly  be  arranged  in  one 
natural  group. 


FIG.  55. — Trypanosomes  of  sleeping  sick- 
ness ( Trypanosoma  gambiense) :  the 
form  on  the  extreme  right  is  in  process 
of  division. 


THE   PATHOGENIC   PROTOZOA  143 

chsetes  are  not  yet  clearly  determined.  These  questions, 
which  seemed  of  zoological  and  philosophical  interest  rather 
than  medical,  have  nevertheless  come  into  the  domain  of 
medicine  since  the  discovery  of  the  microbe  of  syphilis  and  its 
treatment.  From  the  study  of  the  pathogenic  protozoa  there 
have  arisen  many  new  ideas. 

In  the  study  of  bacteria  the  methods  are  their  isolation  and 
pure  culture,  the  study  of  their  biochemical  reactions,  and  of 
experimental  inoculations.  Their  physiology,  i.e.,  the  study  of 
their  functions,  takes  precedence  of  the  study  of  their  forms, 
i.e.,  their  morphology.  In  the  case  of  the  protozoa,  morphology 
has  the  first  place.  It  is  no  longer  a  question  of  describing 
one  cell,  but  a  cycle  of  very  dissimilar  cellular  forms  with  repro- 
ductive phases,  sometimes  sexual,  sometimes  asexual.  And  it 
is  only  at  this  price  that  certainty  can  be  attained  on  the 
methods  of  transmission  of  these  microbes  and  on  the  basic 
ideas  for  medicine  and  hygiene.  Cultivation  has  been  success- 
fully accomplished  only  in  the  case  of  certain  species,  the 
trypanosomes  of  the  rat,  amoebae  in  mixed  culture,  and 
piroplasma,  and  it  does  not  give  the  same  help  as  do  bacterial 
cultivations. 

No  intracellular  protozoon  nas  yet  been  cultivated. 

It  is  the  knowledge  of  the  life-cycle  which  gives  the  key  to 
the  transmission.  The  most  complicated  modes  of  transmis- 
sion among  the  bacteria  are  very  simple  in  comparison  with 
those  of  malaria  and  sleeping  sickness.  The  living  carriers  of 
certain  bacteria,  e.g.,  the  rat-flea  in  the  case  of  plague,  appear 
to  be  carriers  pure  and  simple,  i.e.,  possess  practically  the  same 
importance  as  the  needle  of  a  syringe  or  a  lancet.  The  mos- 
quito is  more  than  a  mere  carrier  of  the  haematozoon  of 
Laveran  ;  it  is  a  second  host  and  in  it,  and  it  alone,  the  parasite 
accomplishes  the  sexual  phase  of  its  life-cycle. 

It  was  originally  thought  that  the  tsetse  fly  ( Glossina  palpalis] 
whose  bite  produces  sleeping  sickness  was  a  simple  carrier  of 
the  virus  and  only  remained  infective  for  a  few  hours  after 
sucking  it  from  the  patient's  body.  But  the  recent  experiments 
of  Kleine,  confirmed  by  Bruce,  have  proved  that  the  tsetse  is 


144. 


MICROBES   AND  TOXINS 


really  a  second  host.  It  remains  infective  for  about  24  to  48 
hours  after  the  moment  of  drawing  infected  blood,  then  for  a 
period  of  about  17  days  it  is  non-infective,  again  becoming 
infective  for  a  period  of  about  60  days'  duration.  The  trypano- 

some  undergoes  in  it 
an  evolution  with 
sexual  reproduction. 

The  protoplasm  of 
protozoa  appears  to 
possess  faculties  of 
adaptation  and  varia- 
tion much  more  exten- 
sive than  that  of  bac- 
teria. Biologically 
speaking  it  is  a  long 
journey  for  a  parasite 
of  the  stomach  of  the 
mosquito  to  reach  the 
blood  of  man.  In  the 
successive  phases  and 
habitats  there  are 
forms  and  structures 
so  different  that  it  re- 
quires strict  proofs  to 
convince  one  that  it  is 
really  the  same  species. 
The  Leishmania  donovani  of  Kala-azar  (a  disease  of  India  and 
the  Mediterranean)  is  in  the  intestine  of  a  bug  a  flagellate 
form  :  in  man  it  is  an  intracellular  form  deprived  of  all 
locomotory  organs.  The  degradation  resulting  from  para- 
sitism has  abolished  in  many  forms  the  characteristic 
structures  and  in  many  cases  permits  of  only  provisional 
classification. 

These  degradation  phenomena  are  more  striking  among  the 
protozoa  because  we  know  on  the  other  hand  their  complete 
cycle.  The  bacteria,  as  we  have  seen,  have  reached  in  their 
apparent  simplicity  perhaps  the  final  stage  of  degradation,  and 


FlG.  56. — Glossina  palpalis,  the  tsetse  fly 
which  carries  sleeping-sickness. 


THE   PATHOGENIC   PROTOZOA  145 

they  conceal  their  origins,  of  which  we  can  with  difficulty 
discover  a  few  vestiges. 

The  protozoa  may  injure  their  host  both  by  mechanical 
and  by  chemical  means.  The  Entamceba  histolytica  destroys 
and  strips  the  epithelial  cells  from  the  intestine,  abolishing 
at  certain  points  the  impermeability  of  the  healthy  mucosa. 
Myxobolus  pfeifferi  produces  atrophy  of  the  muscle  fibres. 
Lentospora  destroys  the  bones  and  cartilages  of  the  trout. 

The  toxins  of  the  protozoa  are  little  known.  If  they  exist 
they  are  difficult  to  isolate  and  demonstrate.  The  Sarcocystine 
of  Laveran  and  Mesnil,  which  kills  the  rabbit  and  only  the 
rabbit,  is  a  well-characterised  toxin,  but  no  similar  substance 
has  been  isolated  from  the  cultures  of  trypanosomes,  or  from 
the  blood  of  animals  with  a  trypanosome  infection.  The  blood 
of  a  malarial  patient,  filtered  at  the  moment  of  the  paroxysm,  is 
not  quite  harmless  to  a  healthy  individual.  The  blood  of  an 
animal  infected  with  trypanosoma  gambiense  produces  an 
appearance  of  somnolence  in  experimental  animals,  but  it  is 
difficult  to  say  how  much  of  this  is  due  to  products  of  the 
parasite  and  how  much  to  the  host.  These  are  researches 
which  will  have  to  be  continued ;  there  is  no  reason  to  believe 
in  advance  that  toxins  and  endotoxins  do  not  exist  in  the 
pathogenic  protozoa. 

Heredity  in  Protozoal  Diseases. — The  protozoa  are 
frequently  intracellular  parasites.  Bacteria  also  may  inhabit 
cells,  for  example  the  bacillus  of  leprosy,  the  bacillus  of  swine- 
erysipelas,  and  the  tubercle  bacillus.  But  in  these  cases  it  is 
the  cell  which  has  taken  up  the  bacterium,  the  cell  being 
mesodermic  and  naturally  phagocytic ;  the  microbe  has  been 
captured ;  no  bacterium  ever  penetrates  by  its  own  activity 
into  a  living  cell.  On  the  contrary,  many  protozoa  have  during 
their  life-cycle  a  motile  form,  amoeboid  or  flagellate,  thanks  to 
which  they  can  penetrate  spontaneously  the  cells  of  their 
host. 

This  fact  is  of  capital  importance  from  the  point  of  view  of 
heredity  in  disease.  When  one  sees  the  young  of  an  anthrax- 
infected  mother  born  with  an  anthrax  pustule,  one  might  think 

L 


146  MICROBES   AND   TOXINS 

that  the  disease  was  hereditary,  but  in  reality  it  is  a  case  of 
contagion  or  of  transmission  at  short  radius ;  the  placental 
filter  has  been  injured  (Chamberland's  experiment).  Nowadays 
heredity  in  tuberculosis  is  no  longer  believed  in;  what  is 
inherited  is  at  most  a  physiological  predisposition  of  the  soil 
(and  even  that  is  a  vague  and  uncertain  idea),  or,  alternatively, 
conditions  of  life  in  which  the  bacillus,  everywhere  to  be 
found,  can  flourish.  There  is  no  hereditary  infection  in  the 
strict  sense  unless  the  fertilised  ovum  is  infected  by  the 


Fig.    57. — The  spirochsete  of  Schaudinn    in    the  liver  of  a  congenital 
syphilitic.     (From  a  microphotograph.     Magnified  3,000  times. ) 

parasite  (among  the  vertebrates  either  from  infection  of  the 
female  cell  or  of  the  spermatozoon,  or  of  both) ;  in  such  a 
case  the  disease  is  truly  congenital.  There  is  no  certain 
example  of  such  a  fact  among  the  bacterial  diseases,  and  this 
is  the  reason  why  the  idea  of  heredity  among  them  has  lost  so 
much  ground. 

From  their  power  of  penetrating  cells  the  protozoa  fre- 
quently infect  parasitically  the  ovum,  thus  producing  hereditary 
infections.  The  first  thoroughly  demonstrated  example  of 
hereditary  infection  was  that  of  the  pttrine  of  the  silk-worm, 


THE   PATHOGENIC   PROTOZOA  147 

rendered  so  famous  by  Pasteur.  Under  the  microscope  the 
presence  of  the  germ  was  demonstrated  in  the  egg,  and  it  was 
recognized  that  the  infected  eggs  produced  caterpillars  which 
formed  the  point  of  departure  for  the  infection  of  the  following 
year.  In  general,**in  this  example  the  heredity  is  only  of  one 
generation,  for  the  silk-worm  infected  in  the  egg  rarely  survives 
to  become  an  adult ;  it  is  the  other  silk-worms,  infected  late  in 
their  larval  stage,  which  succeed  in  reaching  the  adult  con- 
dition after  more  or  less  great  vicissitudes,  which  produce  the 
infected  eggs. 

Eckhardt  has  found  coccidia  (Cocddium  tenellum)  in  the 
white  of  hen's  eggs,  and,  according  to  him,  these  parasites 
produce  an  early  infection  of  the  chicks  which  in  consequence 
very  soon  die. 

The  higher  vertebrates  have  an  interest  in  the  hereditary 
transmission  of  protozoal  diseases  from  two  points  of  view ; 
either  it  occurs  in  themselves  or  it  is  a  condition  of  the 
infection  of  an  invertebrate  host  which  transmits  to  them  the 
disease.  Thus  the  piroplasmosis  due  to  Piroplasma  bigeminum 
is  transmitted  from  ox  to  ox  by  a  tick,  Rhipicephalus  annulatus, 
but  it  is  not  the  same  individual  tick  which  carries  the 
piroplasma  from  one  ox  to  another.  One  tick  becomes 
infected  from  an  infected  ox,  and  it  is  its  progeny,  a  daughter 
tick,  which  infects  the  healthy  animal.  On  the  inheritance  of 

the  parasite  in 
the  insect-carrier 
depends  the  pro- 
pagation of  the 
disease  in  the 
vertebrate. 

FIG.  58.— The  spirochaete  of  syphilis  :  forms  in         An  mhentance 
longitudinal  division.  IS  probable  also  in 

other  diseases  of 

vertebrates  which  are  transmitted  by  ticks,  e.g.,  the  spirillosis  of 
fowls,  African  tick-fever  and  recurrent  fever.  R.  Koch  saw  the 
spirochaete  of  African  recurrent  fever  in  the  egg  of  the 
tick  which  transmits  it,  Ornithodorus  moubata.  In  fowls 

L  2 


148  MICROBES   AND  TOXINS 

infected  with  the  spirochsete  of  Marchoux  and  Salimbeni  the 
parasite  may  penetrate  the  egg,  particularly  the  yolk,  and  in 
this  case  inheritance  may  occur  from  vertebrate  to  vertebrate. 
Hygiene  has  to  take  account  of  these  facts :  to  abolish  a 
parasite  it  is  insufficient  to  destroy  it  in  the  vertebrate ;  the 
invertebrate  host  also  has  to  be  abolished  since  it  is  capable  of 
transmitting  the  parasite  to  its  descendants,  the  egg  of  the 
infected  insect  preserving  the  disease  in  nature  somewhat  as  the 
anthrax  spore  keeps  alive  anthrax. 

Schaudinn  considered  the  spirochaete  which  he  discovered 
in  syphilis  to  be  a  protozoon.  Now  the  clinicians  regard 
syphilis  as  a  disease  which  can  be  inherited.  The  case  of  new- 
born infants  with  syphilis  does  not  alone  prove  an  hereditary 
infection,  the  spirochaete  being  very  motile  might  be  transmitted 
from  the  mother  to  the  foetus  through  some  lesion  of  the 
placenta.  Congenital  syphilis  is  by  no  means  necessarily  a 
syphilis  by  conception.  But  from  certain  observed  facts  such 
true  inheritance  is  very  probable.  The  spirochaete  has  the 
power  of  spontaneously  penetrating  cells :  it  has  even  a 
predilection  for  epithelial  cells :  further,  although  there  are 
no  certain  observations  of  its  presence  in  a  spermatozoon,  it  has 
been  seen  in  the  spermatic  tubules  in  close  relation  to  the 
epithelial  cells  (in  congenitally  syphilitic  boys),  and  it  has  been 
seen  (Levaditi  and  Sauvage)  in  the  protoplasm  of  the  ovarian 
follicles  of  female  children.  Are  infected  ova  in  the  woman 
capable  of  fertilisation,  and,  if  fertilised,  capable  of  normal 
development?  It  seems  possible  in  view  of  the  clinical  facts  and 
by  analogy  with  the  case  of  the  tick  Ornithodorus,  whose  eggs 
infected  with  spirilla  give  rise  to  larvae  which  as  adults  are 
capable  of  conveying  the  infection. 

To  sum  up,  we  know  from  Finger  and  Landsteiner's  experi- 
ments that  the  semen  of  an  adult  syphilitic  is,  as  a  substance, 
capable  of  producing  syphilis,  and  we  know  that  the  congenital 
syphilitic  of  the  male  sex  shows  the  parasite  developing  in  the 
seminal  gland  in  contact  with  the  epithelial  cells,  but  no  one 
has  ever  seen  a  spirochaete  in  an  adult  spermatozoon.  In  the 
female  subject  there  can  be  no  doubt  of  the  possibility  of 


THE   PATHOGENIC   PROTOZOA  149 

transmission  by  the  general  circulation  through  the  blood,  i.e., 
through  the  placenta ;  and  further,  spirochaetes  have  been  seen 
in  the  interior  of  the  ova  in  female  congenital  syphilitics ;  it  is 
not  known  with  absolute  certainty  whether,  and  how,  a  spirochaete 
passes  from  the  general  circulation  of  the  mother,  or  from  the 
spermatic  fluid  of  the  father,  into  the  ovum  which  after  fertilisa- 
tion is  to  produce  an  embryo  infected  from  the  start,  whose 
life  in  consequence  will  be  more  or  less  soon  cut  short.  But 
taking  all  the  facts  we  know  we  have  almost  a  complete 
demonstration  of  true  inheritance. 

Protozoal  disease  and  hereditary  disease  are  two  terms  so 
closely  associated  in  our  minds  to-day  that  the  protozoal  nature 
of  the  spirochaete  is  invoked  to  support  the  hereditary  character 
of  syphilis,  and  this  latter  is  brought  forward  as  an  argument 
in  favour  of  the  protozoal  nature  of  the  spirochaete :  there  are 
at  the  base  of  this  somewhat  easy-going  argument  facts  which 
are  sufficiently  certain. 

The  striking  analogies  between  syphilis,  a  spirochaete  infection, 
and  sleeping  sickness,  a  trypanosome  infection  permit  of  the 
belief  that  the  spirochaete  of  Schaudinn  is  a  protozoon.  Among 
the  more  or  less  late  complications  of  syphilis  are  locomotor 
ataxy  and  general  paralysis.  Now  there  is  also  known  an 
ataxic  condition  in  dogs  infected  with  trypanosomes 
(Spielmeyer's  experiments),  and  there  exists  a  general  paralysis 
with  all  the  mental  stigmata  of  that  disease  in  men  attacked  by 
sleeping  sickness  (G.  Martin  and  Ringenbach). 

There  are  doubtless  more  protozoal  diseases  than  we  think 
to-day,  and  it  may  quite  well  be  that  protozoa  are  the  cause  of 
those  infections  whose  nature  is  still  unknown — e.g.,  yellow 
fever,  cattle-plague,  and  the  horse-sickness  of  the  Transvaal. 
Yellow  fever  in  particular  is  transmitted  by  a  mosquito 
(Stegomyia  fasciata\  which  does  not  infect  until  after  the 
1 2th  day  from  the  time  at  which  it  was  itself  infected.  The 
individual  bitten  passes  through  a  period  of  prostration  which 
lasts  three  to  five  days,  and  at  this  moment  his  blood  becomes 
infective  for  the  mosquito,  but  only  for  a  period  of  three  days ; 
these  facts  indicate  a  life-cycle  in  the  mosquito  and  in  man, 


150  MICROBES   AND   TOXINS 

a  series  of  different  forms  (cf.  malaria)  which  appear  and 
disappear  in  the  blood.  These  forms  are  unknown  and  must 
be  extremely  minute. 

From  the  immunity  point  of  view  also  the  protozoal  diseases 
present  characters  quite  different  from  the  bacterial  infections. 

THE  VIRUSES  CAPABLE  OF  PASSING  FILTERS. 

The  bacteria  which  we  study  under  the  microscope  are 
unequal  in  size.  The  Bacillus  Butschlii  we  mentioned  in 
connection  with  the  nucleus  of  bacteria  is  a  colossus  in  com- 
parison with  the  bacilli  of  fowl-cholera,  with  the  Micrococcus 
parvulus  of  Veillon,  or  even  with  the  little  bacillus  found 
in  influenza  by  Pfeiffer.  There  probably  exist  bacteria  still 
smaller.  Our  best  microscopes  do  not  allow  us  to  dis- 
tinguish a  particle  whose  thickness  is  less  than  o'i  /x.  The 
bacteria  smaller  than  o-i  /x,  are  therefore  invisible  under  the 
microscope  ;  they  are  ultra-microscopic.  Since  there  are  many 
diseases  in  which  the  microbe  remains  unknown  we  are 
tempted  to  ascribe  to  them  ultra-microscopic  microbic  agents. 
Already  in  1884  Pasteur  said  that  the  virus  of  rabies  was  too 
small  for  us  to  be  able  to  see  it. 

The  study  of  these  extremely  small  microbes  only  commenced 
in  1898  with  an  experiment  by  Lofflerand  Frosch  on  the  virus 
of  foot-and-mouth  disease,  which  no  one  has  yet  made  visible. 
The  serous  fluid  from  an  ulcer  (in  which  no  microbe  can  be  seen) 
is  diluted  with  water  and  filtered  through  a  porcelain  bougie 
(similar  to  those  of  the  Chamberland  filters) ;  there  results  a 
perfectly  clear  fluid  free  from  visible  microbes  which  is  capable 
of  transmitting  the  disease  to  a  fresh  animal ;  this  is  the  first 
example  of  a  virus  passing  through  filters,  or,  as  it  is  commonly 
called,  a  filtrable  virus. 

Since  1898,  the  existence  of  filtrable  viruses  has  been  proved 
by  experiments  in  about  twenty  diseases,  the  chief  of  which 
are  foot-and-mouth  disease,  pleuropneumonia  of  cattle  (rinder- 
pest), yellow  fever,  swine-plague,  cattle-plague,  small-pox,  and 
rabies. 


THE   PATHOGENIC   PROTOZOA  151 

The  study  of  these  viruses  is  far  from  being  advanced  ;  one 
only,  that  of  cattle  pleuropneumonia,  has  been  seen  (and  even  its 
form  is  subject  to  discussion),  obtained  in  pure  cultures  and  treated 
like  an  ordinary  bacterium.  In  vaccinia,  small-pox,  sheep-pox, 
hydrophobia,  trachoma  and  molluscum  contagiosum,  microbes 
have  been  described  but  they  are  still  hypothetical ;  the  proof 
is  still  to  seek. 

The  expression,  "  invisible  microbes,"  originally  employed  to 
describe  these,  has  been  abandoned  as  inexact,  and  they  have 
been  called  the  "  so-called  invisible,"  and  later  the  "  filtrable 
viruses."  Invisible  microbes  are  simply  microbes  which  have 
not  yet  been  seen  ;  for  example,  the  syphilitic  virus  was  classed 
among  them  till  the  day  when  Schaudinn  discovered  the 
Spirochaete.  The  classic  example  of  pleuropneumonia  shows 
that  a  microbe  may  traverse  a  porcelain  filter  without  being 
invisible.  Little  vibrios  and  even  little  protozoa  (Micromonas 
Mesnili,  of  Borrel)  have  been  found  in  water;  these  pass 
through  a  filter  and  can  yet  be  quite  easily  seen.  A  filtrable 
microbe  is  not  necessarily  visible. 

The  name  of  ultra-microscopic  microbes  is  the  most  correct, 
because  many  of  these  micro-organisms,  too  small  to  be  seen 
under  the  microscope,  can  be  studied  with  the  ultra-micro- 
scope. Everyone  has  heard  of  this  improvement,  consisting 
in  examining  the  object  not  as  lighted  from  below  and  seen 
by  transmitted  light,  but  lighted  from  the  side  so  as  to  appear 
as  a  bright  point  or  line  on  a  dark  field.  In  the  observation 
of  microbes  which  are  perfectly  visible  the  ultra-microscope  is 
not  the  instrument  of  choice  for  studying  the  structure ;  a  well- 
stained  preparation  is  still  the  best.  The  ultra-microscope 
furnishes  to  medical  bacteriology  above  all  an  economy  of 
time  and  trouble ;  it  makes  the  finding  of  the  microbes  a 
more  rapid  process,  for  example,  when  there .  are  very  few 
trypanosomes  in  the  blood  or  spirochsetes  in  the  fluid  from  a 
lesion  suspected  of  being  syphilitic;  but  these  are  quite 
visible  microbes. 

Are  the  ultra-microscopic  viruses  always  microbes  ?  May 
it  not  be  a  question,  at  least  in  certain  cases  (as  Beijerinck  has 


152  MICROBES   AND  TOXINS 

suggested  for  the  mosaic  disease  of  the  tobacco  plant),  of  a 
fluid,  living  contagium  which  is  literally  invisible?  This 
hypothesis  of  "  soluble  viruses  "  has  been  put  forward,  but 
hitherto  no  positive  proof  has  been  given. 

The  essential  procedure  in  the  definition  of  the  ultra-micro- 
scopic viruses  is  filtration  ;  it  is  the  current  method  of  isolation. 
The  liquid  containing  the  virus  is  passed  through  a  filter — for 
example,  vaccinal  pulp  rubbed  up  in  water;  the  filtrate 
collected  is  virulent,  and  with  it  attempts  at  cultivation  can  be 
made.  The  filters  employed  are  of  the  well-known  forms ; 
the  majority  are  hollow  bougies,  like  those  which  are  used  for 
filtering  drinking  water ;  they  are  made  of  porcelain  (Chamber- 
land  filter),  of  infusorian  earth  (Berkfeld  filter),  of  asbestos, 
charcoal,  plaster,  etc. 

These  filters  do  not  act  towards  microbes  as  does  a  sieve  used 
to  sift  seeds  of  unequal  size,  or  as  the  metallic  grids  used  for 
separating  sand  of  different  coarseness.  It  would  not  be  right 
to  conceive  the  large  microbes  as  being  kept  back  because  they 
are  bigger  than  the  meshes,  whereas  the  little  ones  pass  easily 
through,  just  as  the  smaller  fishes  pass  through  the  meshes  of 
a  net.  Even  the  bacteria  of  average  size,  such  as  the  vibrio  of 
cholera,  are  smaller  than  the  pores  of  our  filters,  and  their  size 
would  permit  them,  to  use  the  simile  of  Duclaux,  to  pass 
through,  as  a  train  passes  through  a  tunnel,  without  rubbing 
against  the  walls ;  what  keeps  them  back  is  that  they  are  held 
against  the  walls  by  the  capillary  pressure. 

Filtration  is  not  a  simple  mechanical  operation,  various 
factors  act  in  it :  the  quantity  of  the  virus,  the  motility  of  the 
microbe,  the  pressure,  the  degree  of  dilution,  the  nature  of  the 
liquid  more  or  less  albuminous,  the  temperature,  the  duration 
of  the  filtration,  and  the  texture  of  the  filter.  All  these  factors 
have  to  be  taken  into  account  in  these  experiments.  As  a  rule, 
several  nitrations  are  performed  one  after  another,  the  first, 
rougher  and  more  rapid,  prepare  for  the  final  one  by  freeing 
the  liquid  from  particles  which  block  up  the  pores.  One  must 
especially  avoid  having  thick  albuminoid  substances  present ; 
they  soon  cover  over  the  surface  of  the  filters.  By  prolonging 


THE   PATHOGENIC   PROTOZOA  153 

the  time  it  would  be  possible  to  make  microbes  pass  which  are 
not  ordinarily  "  filtrable,"  but  this  would  not  be  really  a  filtration 
but  a  culture  propagating  itself  by  extension  from  one  side  of  the 
filter  to  the  other.  This  is  what  happens  in  the  filters  of  water 
supplies  which  are  badly  kept.  Bacteriologically  speaking,  they 
are  no  longer  niters  at  all ;  instead  they  are  continually  infecting 
the  drinking  water  with  the  microbes  which  they  are  supposed 
to  be  keeping  out. 

Filtration  can  show  that  some  quite  minute  microbe  exists 
in  a  given  infection  :  it  gives  no  information  as  to  its  nature. 
Fortunately,  the  labours  of  Jenner  and  Pasteur  have  proved  that 
it  is  possible  to  study  a  virus  without  seeing  it.  It  can  be 
purified  (precisely  by  filters  as  it  happens),  inoculated,  and  its 
resistance  to  physical  and  chemical  agents  (heat,  antiseptics, 
etc.)  determined,  as  well  as  the  conditions  of  preservation  and 
attenuation.  All  the  viruses  enumerated  at  the  head  of  the 
chapter  have  been  treated  in  this  way  and  processes  of  immun- 
ization have  been  discovered  against  certain  of  these  viruses 
which  are  still  unknown,  a  paradox  which  has  become  familiar 
to  us  through  Jennerian  vaccination  and  the.antirabic  treatment 
of  Pasteur. 

To  say  that  a  virus  is  filtrable  is  to  give  it  an  external  rough 
definition ;  there  are  undoubtedly  in  this  group  very  different 
microbes ;  some  may  be  bacteria,  others  protozoa.  Borrel  has 
described  a  protozoon  which  passes  the  rough  filters.  In  the 
life  cycle  of  the  Hamamceba  Ziemanni  of  the  little  owl, 
Schaudinn  has  described  motile  forms  smaller  than  the 
microbe  of  peripneumonia ;  it  is  admitted  that  even  the  most 
visible  protozoon  may  have  ultra-microscopic  stages. 

Several  groups  may  henceforth  be  distinguished  among  the 
diseases  due  to  filtrable  viruses : 

i.  Pleuropneumonia  of  cattle  :  in  this  the  microbe  has  been 
filtered,  cultivated,  and  finally  seen.  It  would  seem  that  it 
ought  to  be  easy  to  describe  it.  At  first  it  was  said  to  be  a 
bacterium,  extremely  fine  cocci  namely,  which  it  was  possible 
to  see  under  the  ordinary  microscope,  not  singly,  but  in 
amorphous  masses.  Recently,  Bordet  has  described  by  means 


154 


MICROBES   AND   TOXINS 


of  culture  in  a  special  medium,  forms  resembling  spirochaetes 
Borrel,  using  a  different  technique,  has  criticised  the  forms 
seen  by  Bordet  and  concluded  that  it  is  not  a  spirochaete  but  a 
new  type  lying  perhaps  between  the  protozoa  and  the  bacteria, 
and  still  incapable  of  precise  definition. 

2.  Cases  of  blood  infection  or  septicaemias  such  as  the 
horse-sickness  and  the  catarrhal  malarial  fever  of  sheep 
(studied  in  particular  in  the  Transvaal),  yellow  fever,  cattle- 
plague,  avian-plague,  and  hog-cholera  appear  to  be  of  the  same 


)> 


oi 


0     * 


FIG.  59. — Various  forms  of  the 
microbe  of  bovine  pleuro- 
pneumonia  according  to  Bordet. 


FlG.  60. — Various  forms  of  the 
microbe  of  bovine  pleuro- 
pneumonia  according  to  Borrel 
(higher  magnification  than  in 
Fig.  59). 


nature.  Horse-sickness  and  the  sheep  disease  which  resembles 
malaria  only  exist  in  localities  where  there  are  certain  definite 
mosquitos ;  like,  malaria,  too  they  were  formerly  called  mias- 
matic disease.  Horses  do  not  take  horse-sickness  even  when 
they  are  exposed  to  conditions  of  climate  and  altitude  reputed 
to  be  dangerous,  provided  they  are  protected  from  mosquitos 
by  means  of  wire-screens.  The  "  heart-water  "  of  ruminants  is 
transmitted  by  a  tick  (the  bont-tick — Amblyomma  hebraeum). 


THE  PATHOGENIC  PROTOZOA  155 

Yellow  fever  is  inoculated  by  a  mosquito,  Stegomyia  fasdata. 
These  diseases  have  all  the  behaviour  of  protozoal  infections. 
By  reason  of  certain  procedures  of  immunization  which  are 
common  to  them  all,  horse-sickness,  cattle-plague,  and  hog- 
cholera  present  some  points  of  similarity. 

3.  The  diseases  characterized  by  localization  to,  and  lesions 
in,  the  epithelium,  for  example,  small-pox  and  vaccinia,  foot- 
and-mouth  disease,  molluscum  contagiosum  of  birds  and  man, 
scarlatina,  the  jaundice  of  silk-worms,  and  trachoma  or  granular 
conjunctivitis  are  perhaps  to  be  classed  in  this  group. 

Small-pox  or  vaccinia  is  the  type-specimen  of  these  infections ; 
the  characteristic  lesion  is  the  pustule  and  the  pustule  is  a 
collection  of  epithelial  cells  containing  the  virus  and  forming  a 
focus  of  culture  in  vivo.  The  cells  which  build  up  the  little 
tumour  have  no  longer  the  normal  structure  of  the  epidermic 
or  Malpighian  cells ;  they  have  become  globular,  voluminous 
and  "  dropsical " ;  the  nucleus  is  altered,  being  swollen  and 
frequently  out  of  position ;  beside  the  nucleus  there  appears  a 
mass  of  abnormal  material  called  the  "  cellular  inclusion." 

This  mass  was  for  long  regarded  as  a  parasite,  as  the  visible 
stage  of  a  protozoon  which  possessed  other  stages  more  minute 
or  ultra  -  micro- 
scopic. Nowadays 
it  is  known  that  it 
is  merely  a  de- 
formation of  the 
nucleus  appearing 

after  the  invasion  FIG.  61.— Negri  bodies  in  rabies;  the  hypothetical 
of  the  cell  by  the  microbe  of  rabies  at  different  stages.  (After 

virus.     It  is,  as  it 

were,  the  stigma  of  the  presence  of  a  virus  within  the  cell,  or 

even  within  the  nucleus. 

These  stigmata  have  been  described  under  different  aspects 
and  names  in  small-pox,  vaccinia  (Guarnieri  bodies),  rabies 
(Negri  bodies),  molluscum  contagiosum  (described  by  Virchow), 
trachoma,  and  the  jaundice  of  the  silk-worm,  and  the  similarity 
of  these  infections  can  no  longer  be  doubted, 


156 


MICROBES   AND  TOXINS 


FIG.  62. — Mallory  bodies;  the  hypothetical 
microbe  of  scarlatina  under  different 
aspects.  (After  Calkins. ) 


Borrel  has  discovered  in  the  cells  of  molluscum  contagiosum 
minute  corpuscles  very  equal  in  size  and  distinct  from  the 
nucleus,  from  the  chromatin  and  from  the  protoplasm :  they 
are  small  enough  to  pass  through  niters,  and  sufficiently 
abundant  and  resistant  to  physical  influences,  as  temperature 

and  drying,  to  explain 
•M  the  powerful  nature  of 

the  contagium  in  these 
diseases. 

This  may  perhaps  be 
the  type  of  microbe  so 
long  sought  for  in 
small-pox  and  granular 
conjunctivitis,  but  it  is 
necessary  to  speak  with 
reserve  as  cultivation 
has  not  yet  been  suc- 
cessful. What  is  certain  is  that  there  exist  ultra-microscopic 
bacteria  and  protozoa  sufficiently  small  to  traverse  the  pores  of 
niters  made  of  asbestos,  porcelain,  plaster,  or  infusorian  earth. 
Great  discoveries  are  still  to  be  made  in  this  domain — a  domain 
opened  up  twelve  years  ago  by  the  study  of  foot-and-mouth 
disease  and  pleuro-pneumonia. 

The  curiosity  of  investigators  ought  not  to  be  monopolised 
by  the  diseases  occurring  in  man  and  animals.  There  is  no 
reason  why  there  should  not  be  invisible  microbes  elsewhere 
in  those  fermentations  which  go  on  everywhere  in  nature. 
They  may  also  quite  well  play  a  part  in  the  life-cycle  of  plants. 
Just  as  insects  produce  injuries  and  mutilations  in  plants,  so 
the  ultra-microscopic  microbes  may  be  responsible  for  the 
variations  and  mutations  which  occur  in  the  vegetable  world. 
Microbiology  may  hope  here  again  to  bring  its  support  to  the 
Darwinian  doctrine. 


CHAPTER   VIII 

THE   TOXINS 

MlCROBIAL   AND   VEGETABLE   TOXINS — ENDOTOXINS 

Microbial  and  vegetable  toxins— Definition — Soluble  toxins — Characters 
— Toxins  and  diastases :  resemblances  and  differences — Incubation 
Penetration  of  the  body— Elective  fixation — Wassermann's  experi- 
ment— Vegetable  toxins  :  ricin,  abrin,  crotin — Production  of 
antitoxins — Endotoxins — Definition — Toxity  of  microbial  bodies — 
Toxin  and  endotoxin  of  the  cholera  vibrio — Do  anti-endotoxins 
exist  ? — Importance  of  intravenous  inoculations. 

MICROBIAL  AND  VEGETABLE  TOXINS 

WE  know  toxins  as  properties,  not  as  substances,  properties 
of  certain  broth-cultures,  or  properties  of  the  bodies  and 
extracts  of  the  bodies  of  bacteria.  Their  nature  and  their 
exact  chemical  constitution  are  unknown,  for  they  are  bound 
up  with  albuminoid  substances  the  chemistry  of  which  is  still 
in  its  infancy. 

The  science  of  toxins  is  therefore  more  physiological  than 
chemical,  and  the  chief  method  of  experimentation  is  on  the 
living  body.  The  quantitative  element  is  introduced  by  mea- 
suring the  incubation  times,  the  temperature,  or  the  magnitude 
of  the  local  phenomena,  such  as  oedema  and  the  duration  of  the 
symptoms  of  intoxication. 

In  certain  cases  it  has  been  possible  to  replace  the  animal 
experiment  by  experiments  in  vitro,  and  to  measure,  with 
exactitude,  certain  phenomena,  easy  to  observe,  such  as  the 
lysis  of  the  red  corpuscles  of  the  blood  (haemolysis). 

Useful  discoveries  are  much  oftener  reached  by  instinct  than 

157 


158  MICROBES   AND  TOXINS 

by  reason.  If  progress  went  on  logical  lines  the  idea  of  anti- 
bodies in  general  ought  to  have  been  the  first,  and  from  it  the 
existence  of  antitoxins  ought  to  have  been  deduced.  But,  on 
the  contrary,  it  was  the  discovery  of  a  particular  antitoxin  which 
led  to  the  study  of  antibodies  in  general. 

In  experimental  medicine,  the  chief  business  is  not  to  build 
up  systems  of  ideas,  i.e.t  to  philosophize,  but  simply  to  search 
patiently  with  many  trials  and  many  a  re-beginning.  The 
inquisitive,  prying,  intuitive  people  have  the  advantage  over 
those  who  reason. 

At  the  very  beginning  of  the  researches  on  toxins  we  find  an 
experiment  of  Pasteur  :  the  filtrate  of  a  culture  of  fowl  cholera 
produced  in  a  fowl  the  symptoms  of  the  disease  in  the  absence 
of  microbes.  At  first  it  was  thought  that  the  bacterial  toxins 
belonged  to  the  group  of  alkaloid  substances,  the  ptomaines, 
found  by  Selmi  in  dead  bodies,  in  certain  molluscs,  and  in 
bacterial  cultures  (e.g.,  muscarine,  neurine,  &c.).  It  is  true  that 
bacteria  can  produce  poisons  of  this  type  (Brieger),  but  these 
poisons  do  not  produce  a  specific  intoxication  like  that 
observed  in  such  a  well-defined  disease  as  tetanus.  Later,  when 
it  had  been  observed  that  microbes  killed  by  heat  are  not 
harmless,  but  when  inoculated  produce  a  local  suppuration, 
attempts  were  made  to  isolate  the  "  poison  "  by  making  protein 
extracts  of  the  bacterial  bodies  ;  but  the  bacterial  proteins 
of  H.  Buchner  are  not  specific  poisons ;  from  very  diverse 
bacteria  one  can  extract  almost  the  same  poisons.  They  con- 
sist of  excretions  or  residues  of  nutrition  of  the  bacteria,  and 
are  found  in  particular  in  old  cultures. 

Excluding  these  alkaloids  and  proteins,  the  following  are  the 
substances  studied  as  toxins  : — 

1.  The  Soluble  Toxins. — The  type-specimen  is  the  diphtheria 
toxin  or  the  tetanus  toxin.     These  are  secretions  of  bacterial 
cells,  just  as  the  pancreatic  juice  is  a  secretion  of  the  gland 
cells. 

2.  The  Endotoxins. — Examples  :    typhoid  endotoxin,  plague 
endotoxin.     These  are  poisons  which  remain  attached  to  the 
cellular  protoplasm,  and  do  not  diffuse  at  all  or  very  little  in 


THE  TOXINS  159 

broth-cultures.  It  is  necessary  to  destroy  the  cell  to  set  free 
the  poison.  The  process  may  be  exemplified  by  the  zymase 
production  of  Ed.  Buchner,  in  which  the  yeast  cell  is  ground 
up  and  expressed. 

3.  A  special  group  has  to  be  made  for  the  poison  of  the 
tubercle  bacillus  (tuberculin),  and  that  of  the  bacillus  of 
glanders  (mallein).  These  diffuse  into  the  broth,  but  there 
remains  some  in  the  substance  of  the  bacterium  ;  hence  the 
bodies  of  tubercle  bacilli  form  an  active  tuberculin. 

Besides  the  specific  toxins,  cultures  may  contain  non-specific 
poisons,  the  proteins  of  Buchner. 

Soluble  toxins.  — The  toxins  of  diphtheria,  of  tetanus,  and  of 
botulismus  are  known  (the  latter  being  produced  by  an 
anaerobic  bacillus  growing  in  meat).  They  can  be  obtained  by 
cultivating  the  bacteria  with  an  abundant  supply  of  air  (in  the 
case  of  the  bacillus  of  diphtheria),  or  in  broth  deprived  of 
oxygen  (in  the  case  of  the  tetanus  bacillus).  The  bacteria  are 
removed  from  the  fluid  by  filtering  through  porcelain  bougies. 
Those  strains  of  bacilli  are  picked  out  which  furnish  the  best 
toxins,  for  good  toxins  are  necessary  for  the  production  of 
good  antitoxins. 

The  Characters  of  the  Soluble  Toxins.— The  soluble 
toxins  possess  very  definite  characters ;  in  the  first  place  they 
are  strictly  specific  ;  the  symptoms  produced  by  the  diphtheria 
toxin  are  quite  distinct  from  the  symptoms  of  tetanus.  Their 
action  is  specific  in  another  sense,  as  it  varies  in  the  different 
animal  species.  The  fowl  does  not  react  to  the  tetanus  toxin 
as  does  the  mouse  or  the  human  subject.  Secondly,  they  are 
extremely  potent :  a  good  culture  of  tetanus  can  kill  the 
mouse  in  a  dose  of  I^-GVTTG  c-m-  The  diphtheria  toxin  kills 
a  guinea-pig  of  250  grams  in  a  dose  of  10400  c.c.1  The 

1  "  I  c.c.  of  the  active  fluid  toxin  produces  on  evaporation  in  vacua  O'OI 
gram  of  dry  residue.  Deducting  the  weight  of  ash  and  the  portion  in- 
soluble in  alcohol  (which  has  no  toxic  activity)  there  remains  0*0004  gram 
of  organic  matter.  It  is  certain  that  the  greater  portion  of  this  weight 
consists  of  substances  other  than  the  diphtheria  toxin.  Yet  this  minute 
quantity  is  sufficient  to  kill  at  least  eight  guinea-pigs  of  600  grams  each, 
or  two  rabbits  of  3  kil."  (Roux  and  Yersin.) — I  c.c.  of  a  good  tetanus 
filtrate  dried  in  vacuo  leaves  0*04  gram  of  dried  residue  of  which  0*025 


160  MICROBES   AND  TOXINS 

botulismus  toxin  is  fatal  in  the  dose  of  Yjnj-^  c-c-  (by  sub- 
cutaneous inoculation  in  each  of  these  cases).  Toxins  are 
soluble  in  water  and  in  glycerine,  and  can  be  precipitated  from 
solution  in  virtue  of  the  fact  that  they  adhere  to  precipitates 
and  coagula ;  precipitation  is  a  method  of  purification.  They 
are  unstable  or  "labile"  substances,  heat,  light,  and  oxygen 
destroying  them  fairly  quickly.  Exposed  for  an  hour  and 
a  half  to  the  temperature  of  55°  C,  the  tetanus  toxin  loses  its 
toxic  properties  ;  at  60°  C.  it  is  destroyed  in  thirty  minutes  ; 
at  68°  C.,  in  five  minutes.  Toxins  bound  up  with  dried 
precipitates  are  more  resistant ;  the  dry  tetanus  toxin  is  still 
slightly  active  after  an  hour  at  80°  C.,  and  even  after  fifteen 
minutes  at  120°  C.  Sunlight  "  inactivates''  a  solution  of  tetanus 
toxin  in  fifteen  to  eighteen  hours.  When  a  photo-dynamic 
substance  (e.g.,  i  per  cent,  eosin),  is  added,  the  toxin  is 
" inactivated "  after  six  hours' exposure  to  light;  with  2*5  to 
5  per  cent,  eosin  it  is  "  inactivated  "  even  in  the  dark. 

Comparison  between  Toxins  and  Diastases. — The 
chemical  constitution  of  diastases  is  not  much  better  known 
than  that  of  the  toxins.  In  comparing  toxins  with  soluble 
ferments  or  diastases,  although  it  is  possible  to  note  analogies,  it 
is  for  obvious  reasons  impossible  to  give  a  chemical  definition. 
Like  diastases,  toxins  act  in  a  very  small  dose,  are  soluble  in 
water  and  in  glycerine,  and  are  weakened  by  filtration,  and  are 
sensitive  to  the  action  of  oxygen,  of  heat,  and  of  light ;  also 
to  changes  in  their  reactions  and  to  various  chemical  substances 
which  "  poison "  or  destroy  them.  Roux  and  Yersin,  who 
pointed  out  these  analogies  at  the  time  of  their  work  on  toxins 
in  1889,  did  not  see  in  them  more  than  a  suggestion:  "It 
seems  to  us  that  the  diphtheria  poison  has  many  analogies 
with  the  diastases.  Its  activity  is  quite  comparable  to  these 
and  to  the  activity  of  venoms.  We  do  not  mean,  how- 
ever, that  it  produces  phenomena  of  hydrolysis  such  as  the 
diastases  produce.  It  neither  inverts  sugar  nor  digests 

gram  is  organic  matter.  Supposing  that  the  whole  of  this  is  toxin  (which 
is  a  great  exaggeration),  the  lethal  dose  for  a  mouse  would  be  o '000,000, 25 
gram. 


THE   TOXINS  161 

fibrin.  If  we  compare  it  to  the  diastases,  we  do  so  without 
forming  any  rash  opinion  as  to  its  chemical  action  and 
simply  in  order  to  sum  up  some  of  its  properties."  This 
reservation  still  holds  good. 

The  disappearance  of  the  toxic  property  by  heating  to  60°  does 
not  necessarily  mean  the  destruction  of  a  diastase.  Heating 
modifies  the  reaction  of  the  fluids,  especially  of  organic  fluids, 
and  coagulates  the  proteins  :  this  coagulation  may  inhibit  the 
action  of  certain  substances  or  properties  without  destroying 
them.  The  toxins  have  a  character  possessed  neither  by 
chemical  poisons,  e.g.,  strychnine  or  potassium  cyanide,  nor  by 
the  diastases  :  the  action  of  these  chemical  poisons  is  almost 
instantaneous,  and  a  zymase  put  into  a  solution  of  sugar  begins  to 
ferment  it  at  once.  The  toxins,  on  the  other  hand,  when  injected 
into  the  body,  only  manifest  themselves  after  a  silent  period  of 
apparent  inactivity,  the  period  of  incubation.  By  changing 
the  path  of  introduction,  e.g.,  by  injecting  intravenously  instead 
of  subcutaneously,  and  by  increasing  the  dose,  the  period  of 
incubation  may  be  rendered  shorter  :  it  is  never  reduced  to 
zero.  Further  the  minimum  incubation  period  varies  with  the 
species  of  animal  inoculated. 

It  is  admitted  that  the  tetanus  poison,  to  reach  the  nerve- 
centres,  has  to  travel  along  the  peripheral  nerves  from  the  site 
of  its  production,  and  within  certain  limits  the  period  of 
incubation  varies  in  proportion  to  the  distance  of  this  :  but  even 
when  this  delay  is  cut  out  the  incubation  period  does  not 
reach  zero.  Meyer  and  Ransom  inoculated  the  nerve-centres 
of  cats  directly  and  still  found  a  minimum  incubation  of  three 
to  five  hours. 

This  inevitable  incubation  period  suggests  that  the  toxin  of 
the  culture  is  not  the  poison  which  kills  the  animal,  but  that 
the  toxin  inoculated  undergoes  in  the  body  certain  transforma- 
tions (fermentations?)  which  produce  the  true  poison,  the 
action  of  which  is  direct  and  immediate.  This  secondary  poison 
was  said  to  have  been  demonstrated  by  inoculating  mice  with 
extracts  of  the  organs  of  animals  actually  in  tetanus,  but  these 
experiments  have  not  given  constant  results  as  the  symptoms 


162  MICROBES   AND  TOXINS 

produced  differed  from  the  pure  tetanic  symptoms.  It  is  not 
yet  possible  to  believe  in  the  existence  of  pro-toxins  analogous 
to  the  pro-diastases  such  as  the  pro-fibrin-ferment. 

The  toxins  act  in  extremely  minute  doses,  like  the  diastases, 
of  which  a  very  small  quantity  can  determine  a  chemical 
change  in  a  very  large  mass  of  material.  But  there  are 
alkaloid  poisons  which  also  act  in  a  very  small  dose  :  a  man 
dies  after  absorbing  the  five-millionth  part  of  his  weight  of 
aconitine  or  digitalin.  The  mere  fact  of  possessing  an  action 
produced  by  very  small  doses  is  not  equivalent  to  being  a 
diastase  ;  it  may  be  an  ordinary  chemical  phenomenon  ;  an 
artificial  catalyst  such  as  colloidal  platinum  is  "  inactivated  " 
by  one  thousand-millionth  part  of  hydrocyanic  acid;  Graham's 
solution  of  ferric  hydrate  under  certain  conditions  is  sensitive 
to  the  presence  of  i/5,ooo,ooothof  ferrocyanide  of  potassium.1 
Finally,  the  phenomena  produced  by  toxins  are  phenomena 
taking  place  in  living  creatures,  which  makes  it  all  the  more 
difficult  to  determine  whether  they  are  diastasic  in  nature  or 
simply  due  to  ordinary  chemical  reactions.  It  is  true  that  the 
toxins  of  diphtheria  and  tetanus  can  "kill"  20  to  100 
million  times  their  weight  of  living  animal,  but  these  figures 
must  not  be  allowed  to  produce  this  illusion  :  the  quantitative 
relationship  is  complicated  by  a  qualitative  element  whose 
importance  cannot  be  exaggerated.  When  a  horse  is  killed 
by  i/8o,oooth  of  its  weight  of  tetanus  toxin,  the  toxin  is  not 
acting  on  the  whole  mass  of  the  horse  :  to  produce  death  it 
is  sufficient  for  it  to  act  on  the  medullary  nucleus  of  the 
vagus  nerve,  a  group  of  cells  scarcely  weighing  two  grams. 
The  diphtheria  toxin  similarly  acts  in  an  elective  fashion  on  a 
group  of  cells  in  the  medulla  or  in  the  cardiac  ganglia.  To 
take  facts  of  another  order,  the  fixation  of  carbon  monoxide 
on  the  haemoglobin  of  the  blood  is  not  a  diastasic  phenomenon 
(on  ihe  contrary  it  arrests  a  vital  diastasic  function  or  the 
first  importance),  yet  the  toxic  power  of  carbon  monoxide  in 
proportion  to  the  weight  of  the  body  is  certainly  more  than 
100,000. 

1  J.  Duclaux — La  chimie  de  la  matiere  vivante,  Chapitre  X. 


THE  TOXINS  163 

Penetration  of  the  Body. — To  reach  the  sensitive  cells 
the  toxins  do  not  always  follow  the  same  path.  Injected 
subcutaneously  they  pass  into  the  lymph,  then  into  the  blood. 
Injected  into  the  blood  they  save  time,  since  the  passage 
through  the  lymph  is  avoided.  Introduced  into  the  alimentary 
canal  the  botulismus  toxin  retains  its  potency,  but  the 
diphtheria  toxin  and  the  tetanus  toxin  are  inactive  even  after 
being  swallowed  in  much  more  than  a  lethal  dose.  This 
destruction  or  neutralization  cannot  be  attributed  to  an  action 
peculiar  to  the  intestinal  epithelium  nor  to  any  extent  to  the 
influence  of  the  bacteria  and  their  fermentations  in  the 
digestive  tube,  but  chiefly  to  the  action  of  the  digestive 
secretions,  the  pepsin,  and  above  all  the  pancreatic  juice. 

According  to  the  experiments  of  Meyer  and  Ransom  and  of 
Marie  and  Morax,  the  tetanus  toxin  does  not  pass  directly  from 
the  site  of  inoculation  to  the  nerve  centres  ;  it  penetrates  the 
peripheral  nerves  at  their  motor  terminations,  and  follows  these 
nerves  to  reach  the  centres.  All  three  species  of  nerve  fibre, 
motor,  sensory,  and  sympathetic,  can  carry  it ;  but  the  carrying 
power  of  the  nerve  depends  absolutely  on  the  integrity  of  the 
axis  cylinder.  By  employing  antitoxin  it  is  possible  to  localize 
the  action  of  the  toxin  to  certain  definite  territories  and  paths. 
Dissociation  experiments  have  shown  that  the  antitoxin  acts 
by  neutralising  the  toxin  still  in  circulation,  but  is  no  longer 
capable  of  neutralising  toxin  absorbed  by  the  nerve  trunks 
(these  do  not  absorb  antitoxin).  The  fact  that  in  certain 
animals,  man  and  the  horse,  tetanus  always  begins  by  a  con- 
traction of  the  muscles  of  the  jaw  ("  lockjaw  ")  only  means 
that  even  after  a  stab  or  a  wound  at  the  end  of  a  limb  sufficient 
toxin  passes  sufficiently  quickly  into  the  circulation  to  affect 
the  centres  on  which  the  innervation  of  these  muscles 
depends. 

The  cells  of  the  central  nervous  system  are  sensitive  to 
many  poisons,  whether  these  reach  them  by  the  nerve  filaments 
or  through  the  blood.  Directly  introduced  into  the  centres 
the  poisons  act  in  a  smaller  dose,  and  often  produce  different 
symptoms  from  poisons  injected  subcutaneously  or  into  the 

M    2 


164  MICROBES  AND  TOXINS 

blood  stream.  Cerebral  tetanus  is  more  like  a  mental  disease, 
a  sort  of  delirium,  than  the  systematic  tetanic  contractions 
which  follow  the  wound  of  a  limb.  In  the  rat  which  has 
received  tetanus  toxin  intracerebrally,  the  incubation  is  from 
forty-eight  hours  to  three  days,  and  if  the  observer  did  not 
know  with  certainty  that  he  had  injected  tetanus  toxin  he 
would  never  recognize  tetanus  in  the  disease  which  he  observes. 
Psychical  manifestations  predominate  ;  the  rat  is  anxious  and 
vigilant;  without  apparent  cause  it  is  seized  with  sudden 
terrors,  and  runs  madly  round  its  cage.  .  .  .  During  the  crisis 
it  seems  to  obey  an  internal  impulse  ....  and  on  careful 
observation  the  question  forces  itself  whether  many  psychical 
phenomena  in  man  may  not  also  be  produced  by  the  fixation 
on  certain  nerve  cells  of  bacterial  toxins  elaborated  in  the 
intestine  or  in  some  other  part  of  the  body  at  some  particular 
moment  (Roux  and  Borrel).1 

Selective  Fixation  of  Toxins. — For  a  toxin  to  kill  in 
the  minimum  dose  it  must  possess  a  selective  affinity  for  cells 
whose  function  is  important,  and  must  proceed  to  fix  itself  on 
these  cells  and  not  on  others,  when  introduced  into  the 
circulation.  Thus  it  is  necessary  for  a  medullary  nucleus  to 
attract  to  itself  the  few  thousandths  of  a  milligram  of  tetanus 
toxin  introduced  into  a  human  body.  The  intoxication 
depends  entirely  on  this  fixation  of  the  toxin,  and  it  has  long 
been  compared  to  a  dyeing  process. 

Even  in  the  inorganic  world,  and  among  dead  substances 
of  animal  or  vegetable  origin,  numerous  examples  exist  of 
the  fixation  of  a  substance  in  solution  more  dilute  even  than 
are  the  toxins  in  the  blood.  The  examples  which  follow  are 
taken  from  the  book  of  J.  Duclaux  already  quoted.  Pre- 

1  In  the  rat  TV  c-c«  of  diphtheria  toxin  subcutaneously  does  not  produce 
even  local  oedema,  but  a  rat  receiving  this  dose  intracerebrally  is  soon 
completely  paralysed,  and  after  two  or  three  days  of  inertia  it  succumbs. 
A  rabbit  of  less  than  1,500  grams  supports  perfectly  30  centigrams  of  a 
morphine  salt  injected  subcutaneously,  whereas  I  milligram  of  morphine 
hydrochloride  injected  into  the  brain  produces  almost  immediate  effects  in 
a  rabbit  of  the  same  weight.  A  tuberculous  guinea-pig  succumbs  when 
injected  intracerebrally  with  a  dose  of  tuberculin  200  times  smaller  than 
when  injected  subcutaneously. 


THE  TOXINS  165 

cipitated  sesquioxide  of  iron  absorbs  arsenious  acid  (and 
probably  also  phosphoric  acid)  till  there  remains  in  the  fluid 
less  than  one-thousand-millionth  part  (A.  Gautier).  A  skein 
of  white  silk  dipped  in  a  solution  of  eosin  so  dilute  that  the 
eye  can  perceive  no  colour,  i.e.t  to  about  one  in  a  million,  is 
dyed  pink  in  the  course  of  a  few  hours.  Silk  can  take  up 
1*3  per  cent,  of  picric  acid  from  a  solution  containing  only 
0-006  per  cent.  The1  absorptive  power  increases  the  greater 
the  dilution.  Passing  to  living  cells,  we  find  that  a  culture  of 
Aspergillus  niger  can  take  up  from  a  solution  measuring 
250  c.c.  and  containing  one-half  milligram  of  zinc,  i.e., 
1/500,000,  practically  the  whole  of  this  metal  (Javillier) ;  the 
proportion  of  zinc  increases  to  about  i  in  10,000  in  the 
aspergillus  cells  and  falls  below  i  in  10,000,000  in  the  fluid 
which  remains.  Certain  plants  can  absorb  copper  from 
solutions  containing  only  i  in  100,000,000.  Certain  marine 
plants,  such  as  the  ordinary  sea-wrack,  fix  abundantly  the 
iodine  and  silver  which  exist  in  traces  only  in  sea-water. 

The  whole  industry  of  dyeing  is  founded  on  similar  fixations : 
fabrics  are  sensitive  to  and  fix  selectively  the  colouring 
materials.  Picric  acid,  which  stains  the  skin,  does  not  dye 
cotton.  The  microscopical  preparations  of  histologists  and 
biologists  depend  on  this  principle  of  the  selective  fixation  of 
the  stains  by  different  anatomical  elements :  magenta  is  fixed 
by  the  nucleus,  picric  acid  by  the  protoplasm,  indigo-carmine 
by  the  connective-tissue  fibres. 

Ehrlich  has  pointed  out  that  in  jaundice  the  kidneys  and  the 
liver  become  charged  with  bile-pigments,  whereas  the  brain 
remains  free.  When  certain  derivatives  of  paraphenylene- 
diamine  are  administered  to  mice,  the  central  part  of  the 
diaphragm  is  found  stained  brown  much  more  intensely  than 
the  periphery,  and  the  muscles  of  the  eyes,  of  the  larynx, 
and  of  the  tongue  are  much  more  deeply  stained  than  other 
muscles :  this  may  be  because  these  muscles  are  in  continual 
activity,  receive  more  blood,  and  are  the  seat  of  more  intense 
oxidations.  Methylene  blue  in  the  living  animal  is  fixed  by 
the  sensory  fibres,  by  the  nerve-endings  for  taste  and  smell, 


166  MICROBES   AND  TOXINS 

by  the  nerves  of  the  plain  muscles  and  the  cardiac  muscle,  and 
by  certain  fibres  in  the  nerve  centres  :  it  does  not  stain  the 
motor-endings  of  striped  muscle  with  the  exception  of  those  of 
the  eyes,  of  the  larynx,  and  of  the  diaphragm  (Ehrlich). 

The  majority  of  the  staining  substances  fixed  by  the  cerebral 
cortex  are  also  taken  up  by  fatty  substances  :  now  the  cortical 
cells  are  rich  in  lipoids  such  as  cholesterin,  lecithin,  and 
cerebrin.  The  substances  which  stain  "  in  vivo "  are  soluble 
in  these  lipoids,  the  non-vital  stains  are  not.  Narcotics, 
cerebral  poisons,  have  an  activity  proportional  to  their 
co-efficient  of  absorption  by  the  lipoids,  and  both  this  activity 
and  this  co  efficient  vary  with  the  temperature.  Antipyretics 
also  have,  without  doubt,  a  selective  action  on  certain  cells. 

The  famous  experiment  of  Wassermann  and  Takaki  showed 
that  a  similar  selective  absorption  takes  place  between  the 
cerebral  grey  matter  of  mammals  and  tetanus  toxin  when  these 
are  mixed  together  in  a  test-tube;  after  a  certain  time  the 
tissue  fixes  the  toxin  and  the  liquid  is  no  longer  toxic. 

The  brain  of  cold-blooded  animals,  e.g.,  the  lizard  and  the 
tortoise,  does  not  fix  toxin  at  all,  or  very  little.  The  brain  of 
the  frog  does  not  fix  toxin  in  vitro  (and  yet  the  frog  kept  in  a 
fairly  hot  room  is  sensitive  to  tetanus  toxin).  The  fixing 
power  of  the  brain  seems  to  be  proportional  to  its  lipoid 
content,  and  there  is  less  of  these  substances  in  the  brain  of 
cold-blooded  animals.  Brain  material  treated  with  ether, 
which  dissolves  the  fats,  loses  a  great  part  of  its  power  of 
fixation :  brain  which  has  been  boiled  no  longer  fixes  at  all. 
Filtration  of  a  suspension  of  brain  material  (removing  the  cell 
elements)  also  destroys  the  fixing  property.  Cholesterin, 
lecithin,  and  cochineal,  a  fatty  material  extracted  from  the 
cochineal  insect,  all  fix  toxin,  but  when  heated  to  60°  in 
presence  of  moisture,  or  after  a  previous  maceration  in  an 
alkaline  fluid,  the  latter  is  no  longer  a  fixative. 

The  fixation  of  toxin  by  sensitive  cells  is  a  phenomenon  of 
the  order  of  dyeing  or  in  vivo  staining — a  phenomenon  of 
molecular  adhesion. 

There  is  not  in  Wassermann's  experiment,  as  was  thought  at 


THE  TOXINS  167 

first,  a  destruction  or  neutralisation  of  the  toxins  by  an  anti- 
toxin elaborated  by  the  brain  cells.  The  fixation  can  be 
modified  by  altering  the  physical  conditions  :  brain  matter 
emulsified  in  physiological  saline  solution  (o'8  per  cent.)  is  a 
stronger  fixative  than  the  same  emulsified  in  distilled  water, 
but  ten  times  weaker  than  the  same  emulsified  in  salt  solution 
of  10  per  cent.  The  brain  gives  up  the  toxin  more  or  less 
quickly  and  restores  the  toxicity  to  water  when  it  is  allowed  to 
macerate,  and  also  after  drying  or  digestion  with  papaine :  the 
toxin  liberated  has  all  the  biological  properties  which  it  had 
before  its  intimate  contact  with  the  brain  matter.  This 
observation  negatives  the  hypothesis  of  a  secondary  toxin 
elaborated  by  the  cells  from  the  toxin  received  and  capable  of 
acting  immediately  without  incubation. 

It  is  because  of  this  fixation  property  of  the  cells  of  the 
body  that  the  toxin  injected  disappears  more  or  less  rapidly 
from  the  blood  and  cannot  be  recovered,  or  only  in  very  small 
degree,  from  the  excretions.  In  the  rabbit  seventeen  hours  after 
injection  no  free  toxin  is  to  be  found  either  in  the  blood  or  in 
the  organs,  and  there  is  never  any  in  the  blood  at  the  moment 
when  the  tetanic  symptoms  commence  (about  forty-eight  hours 
after  intravenous  inoculation. — A.  Marie). 

Since  the  tetanus  toxin  may  be  fixed  on  cells  other  than  the 
nerve-cells,  it  is  evident  that  the  former  keep  back  at  least  a 
portion  of  the  toxin,  acting  thus  as  a  sort  of  screen  to  the 
nerve-cells.  For  example,  the  rabbit  is  less  sensitive  to  tetanus 
than  the  mouse  and  the  guinea-pig  because  its  spleen  fixes  the 
toxin  and  saves  its  brain.  Hence  it  is  not  the  power  of 
fixation  in  general  which  explains  the  sensitiveness  of  a 
particular  animal,  but  the  selective  fixation  on  certain  definite 
cells  whose  activity  is  indispensable  to  life,  for  example,  the 
cells  of  the  medullary  nuclei  or  of  the  sympathetic  ganglia. 

Scorpions  can  stand  very  large  doses  of  tetanus  toxin 
without  symptoms ;  the  toxin  rapidly  disappears  from  the  blood 
and  accumulates  in  the  liver.  The  alligator,  which  is  re- 
fractory to  tetanus,  retains  in  its  blood  for  more  than  a  month 
toxin  which  has  been  injected  into  it.  The  carp,  the  axolotl, 


168  MICROBES   AND  TOXINS 

and  the  frog,  kept  at  a  low  temperature,  do  not  take  tetanus  and 
retain  the  toxin  intact  in  their  blood  for  months.  The  tortoise, 
which  does  not  take  tetanus  either  at  high  or  low  temperatures, 
retains  in  its  blood  for  months  enough  toxin  to  give  tetanus  to 
mice  on  injecting  it.  The  fowl  also,  very  little  sensitive  to 
tetanus,  retains  toxin  in  its  blood  for  long  periods. 

The  frog,  which  is  refractory  to  tetanus  in  winter  or  when 
kept  at  a  low  temperature,  takes  tetanus  in  summer  or  when  it 
is  kept  warm  in  an  incubator  at  about  30°  C.  At  this  tempera- 
ture the  poison  disappears  from  the  blood  and  the  organs  much 
more  quickly  than  in  the  cold.  The  course  of  the  tetanic 
symptoms  can  be  interrupted  at  will  in  the  frog  kept  in  the 
incubator  by  putting  it  again  at  a  low  temperature.  In  this 
way  the  phenomena  may  be  suspended  as  long  as  the  chilling 
continues ;  if  it  is  again  put  in  the  incubator  the  symptoms 
recommence  at  the  stage  at  which  they  were  interrupted.  In 
the  frog,  fixation  and  response  are,  therefore,  to  a  certain 
extent  dissociated,  for  in  the  cold  the  toxin  is  fixed  by  the  cells 
yet  the  disease  does  not  appear. 

Vegetable  Toxins. — The  bacterial  toxins  are  vegetable 
toxins  since  bacteria  are  microscopic  plants.  They  have  their 
analogies  in  the  higher  plants,  for  example,  ricin  extracted  from 
the  seeds  of  the  common  castor  oil  plant ;  abrin  from  the 
Abrus  precatorius  or  jequirity ;  crotin  from  the  plant  croton 
tiglium. 

Ricin  inoculated  subcutaneously  can  kill  a  rabbit  in  a  dose 
of  0*1  mg.  per  kilo,  body-weight;  it  agglutinates  into  masses 
and  dissolves  the  red  corpuscles  of  the  blood.  The  agglutina- 
tion is  so  rapid  and  powerful  that  it  is  necessary  to  keep 
shaking  the  tube  in  order  to  perceive  the  haemolysis.  The 
chemical  nature  of  ricin  is  not  exactly  known ;  it  is  not 
absolutely  certain  that  it  is  an  albuminoid  substance  (Jacoby). 
Abrin  agglutinates  the  red  corpuscles  but  is  not  a  powerful 
lysin.  Crotin  requires  a  dose  of  several  centigrams  per  kilo,  to 
kill  a  rabbit. 

Toxins  and  Antitoxins. — A  fundamental  difference 
exists  between  the  poisons  of  known  chemical  composition, 


THE  TOXINS  169 

such  as  the  alkaloids  and  glucosides,  and  the  toxins  properly 
speaking.  The  toxins  alone  produce  antitoxins  in  the  animal 
treated  by  graduated  doses,  />.,  the  antitoxins  employed  in 
serotherapy.  Antitoxins  exist  against  ricin,  abrin,  and  crotin,  but 
there  are  none  against  solanin  and  saponin,  which  are  glucosides. 
The  glucosides  although  capable  of  being  fixed  by  sensitive 
cells,  do  not  produce  anti-glucosides.  Only  those  poisons 
which  are  capable  of  giving  rise  to  antitoxins  can  be  regarded 
as  true  toxins. 


ENDOTOXINS. 

In  contrast  to  the  soluble  toxins,  the  endotoxins  are  denned 
as  the  poisons  contained  in  the  bodies  of  bacteria  and  not 
spontaneously  set  free  in  cultures.  Whereas  the  toxins  are 
secretions  of  living  bacteria,  the  endotoxins,  according  to  the 
strict  definition  of  R.  Pfeiffer,  are  only  set  free  by  the 
destruction  of  the  bacterium.  It  is  the  protoplasm  of  the 
bacterium  itself  which  acts  as  a  poison  on  absorption  by 
the  body. 

To  bring  the  endotoxins  into  line  with  a  well-known  example, 
one  may  compare  them  to  the  zymase  of  Buchner,  the  ferment 
of  yeast  not  excreted,  or  scarcely  excreted,  but  set  free  by 
grinding  up  with  sand  and  expression  of  the  juice  under  a 
pressure  of  several  hundred  atmospheres.  But  this  com- 
parison is  not  to  be  taken  to  mean  that  the  endotoxins  are 
diastases. 

Bacterial  extracts  have  been  produced  by  Buchner's  process, 
but  nowadays  endotoxins  are  obtained  by  simpler  procedures. 
MacFadyen  grinds  up  the  microbes  at  the  temperature  of 
liquid  air ;  others  subject  the  cultures  to  combined  maceration 
and  shaking.  Besredka  takes  young  cultures,  dries  them,  and 
re-suspends  them  in  saline  solution. 

The  endotoxins  are  distinct  from  the  proteins ;  the  latter  are 
practically  the  same  in  the  different  species  of  bacteria.  To 
deserve  their  name  they  ought  to  be  specific  and  to  give  rise 
to  antitoxins. 


170  MICROBES   AND  TOXINS 

It  is  in  the  cholera  vibrio,  the  plague  bacillus,  the  typhoid 
bacillus,  and  the  bacillus  of  dysentery  that  they  have  been 
chiefly  sought  for  and  thought  to  have  been  found.  It  is 
necessary  to  employ  this  qualified  method  of  expression, 
because  their  definition  and  even  their  existence  is  still 
disputed.  This  chapter  in  the  physiology  of  bacteria  contains 
many  uncertainties,  and  the  facts  observed  cannot  always  be 
made  to  agree. 

The  endotoxins  obtained  by  different  workers  from  the  same 
bacterium  do  not  seem  to  possess  the  same  properties, 
especially  from  the  point  of  view  of  the  toxic  dose  and  the 
resistance  to  temperature ;  but  it  is  also  well  known  that  with 
tetanus  toxin  different  workers  do  not  obtain  equal  specimens 
either. 

Between  the  endotoxins  described  as  having  the  same 
general  characters  there  exist  differences  which  are  not  to  be 
found  between  the  diphtheria  and  tetanus  toxins;  thus  the 
plague  endotoxin  is  destroyed  by  heat  from  about  75°  C. 
onwards,  whereas  the  typhoid  endotoxin  resists  1 2  7°  C.  (accord- 
ing to  Besredka).  But  physical  differences  are  known  to  exist 
between  the  diphtheria  and  tetanus  toxins  also,  which,  though 
less  striking,  are  none  the  less  real. 

The  dysentery  endotoxin  is  easily  obtained,  the  typhoid 
endotoxin  less  easily,  the  cholera  endotoxin  with  great 
difficulty. 

The  following  are  the  principal  points  disputed. 

i.  The  fundamental  phenomenon  on  which  depends  the 
existence  of  specific  endotoxin  is  the  toxicity  of  the  bacterial 
bodies  themselves.  The  nature  of  the  bacterial  bodies  must 
first  be  agreed  upon,  and  no  agreement  will  be  reached  unless 
young  bacteria  are  taken,  avoiding  as  much  as  possible  the 
alterations  caused  by  manipulations,  however  carefully  or 
cautiously  conducted.  Pfeiffer  is  inclined  to  regard  with  some 
suspicion  the  endotoxins  which  accumulate  spontaneously 
in  old  cultures,  considering  them  as  products  of  a  destruction 
of  the  bacteria,  which  is  accompanied  by  chemical  alterations 
(fermentative),  of  the  details  of  which  we  are  ignorant,  These 


THE  TOXINS  171 

errors  may  be  avoided    by    employing  young  bacteria   from 
cultures  of  twelve  to  eighteen  hours  on  solid  media. 

2.  But  in  many  cases  difficulties  are  met  with  when  one 
attempts  to  determine  the  relations  between  this  endotoxin  and 
the   soluble    poisons   secreted    by    the    same    bacteria    and 
apparently  true  toxins.      For  example,  the  cholera  vibrio  cer- 
tainly contains  an  endotoxin  in  Pfeiffer's  signification,  but  it  is 
none  the  less  true  that  the  cultures  contain  an  excreted  poison, 
the  poison   studied  by  Roux,  Metchnikoff,  and  Salimbeni,  on 
which  the  hope  of  a  serotherapy  in  cholera  has  been  founded. 
It  is  this  poison  which  is  absorbed  into  the  body  and  produces 
the  cramps,  the  chilling,  and  death,  whereas  the  vibrios,  however 
numerous  they  may  be,  remain  in   the  intestine  and  only  very 
rarely  invade  the  blood  and  the  tissues.     "  The  free  poison,'' 
says  Pfeiffer,  "  does  not  exist  in  cultures  except  when  these  are 
already  old  and  contain  many  vibrios  already  destroyed  and 
autolysed."      The  cholera  toxin  of  Roux  and   Metchnikoff  is, 
however,  chiefly  secreted  during  the  first  days  of  a  culture 
kept   under  the  requisite  conditions  (a  virulent  vibrio  and  a 
well-aerated    culture).      This    example   is    not    in   favour   of 
Pfeiffer's  opinion,  at  least  in  the  case  of  cholera,  for  although 
he  admits  that  the  endotoxic  bacteria  can  also  secrete  other 
soluble  poisons,  he  maintains  that  the  latter  are  quite  different 
from  endotoxins  and  do  not  possess  their  specificity. 

3.  Do  the  endotoxins  give  rise  to  antitoxins  in  the  body  of 
an  immunised  animal,  as  do  the  soluble  toxins  ?     This  is  the 
most  disputed  question  of  all.     Without  denying  the  existence 
of  anti-endotoxins  in  principle,  Pfeiffer's  school  considers  that 
hitherto  none  have  been   obtained  which  have  passed  satis- 
factory tests,  and  that  the  sera  prepared  against  plague  and 
typhoid   fever   have   not  hitherto    been    successful    precisely 
because  they  do  not  contain  anti-endotoxin. 

To  prepare  an  anti-endotoxin,  as  in  the  preparation  of  an 
antitoxin,  it  is  necessary  to  inject  several  times  into  an  animal, 
for  example,  the  horse,  the  toxic  substance,  in  this  case  the 
bacterial  bodies,  entire  or  broken  up.  Good  results  are  not 
obtained  when  endotoxins  are  injected  subcutaneously ;  intra- 


172  MICROBES   AND  TOXINS 

venous  injection  is  essential.  By  intravenous  injection  of  young 
microbes,  Besredka  obtained  anti-endotoxins  which,  both  in 
vitro  and  in  the  animal,  neutralise  the  toxic  action  of  the 
bacterial  bodies.  Since  the  production  of  antitoxin  is  the 
essential  character  of  toxins,  these  experiments  would  prove 
the  reality  of  endotoxins  as  specific  poisons. 

It  is  difficult  to  say  to  what  degree  the  sera  hitherto 
prepared  against  typhoid,  plague,  and  dysentery  are  anti- 
microbic  or  antitoxic,  i.e.,  active  against  infection  or  against 
intoxication. 

Although  inferior  to  antidiphtheritic  or  antitetanic  sera,  the 
antidysenteric  and  antiplague  sera  have  already  given  results 
sufficiently  good  to  encourage  us  to  bring  to  perfection  the 
endotoxins  and  their  anti-endotoxins. 


CHAPTER  IX 
TUBERCULIN  AND  MALLEIN — ANIMAL  TOXINS — VENOMS 

Tuberculin  and  mallein — Koch's  phenomen — Action  of  tuberculin — Local 
and  general  reactions— Resistance  of  tuberculin  towards  physical 
agents  which  destroy  other  toxins — Specificity — No  antituberculin — 
Habituation  to  tuberculin — Cutaneous  reaction  of  v.  Pirquet — Tuber- 
culin and  anaphylaxis. 

Animal  toxins — venoms — The  venoms  in  the  animal  kingdom— Snake 
poisons— Physiological  action  of  venoms — Digestive  properties — 
Haemolysis  by  venoms — Role  of  lecithin — Lecithids — Natural  immunity 
of  certain  animals  towards  venoms. 

TUBERCULIN  AND  MALLEIN 

THESE  poisons  are  found  in  old  broth  cultures  of  the 
bacillus  of  tubercle  and  the  bacillus  of  glanders.  They  are 
prepared  by  combined  maceration  and  heat  from  glycerine 
extracts  of  the  cultures.  They  exist  also  in  the  bodies  of  the 
bacteria  and  are  thus  in  a  sense  endotoxins.  But  hitherto  no 
antitoxins  to  them  are  known. 

Tuberculin  and  mallein  from  their  physiological  properties 
occupy  a  place  apart.  Tuberculin  may  be  taken  as  the 
type. 

Koch's  Phenomenon. — The  discovery  of  tuberculin 
originated  in  the  "  Koch's  phenomenon " :  when  tubercle 
bacilli  are  inoculated  subcutaneously  in  a  guinea-pig  nothing 
is  seen  at  the  point  of  inoculation  for  from  ten  to  fourteen  days, 
then  a  nodule  appears  which  later  produces  an  open  sore, 
which  refuses  to  heal ;  the  corresponding  lymphatic  glands 
are  swollen.  If,  however,  a  guinea-pig  already  tuberculous  is 
reinoculated  after  four  to  six  weeks,  there  appears  on  the  third 

173 


174  MICROBES   AND  TOXINS 

day,  without  any  nodule  developing,  a  necrosis  of  the  skin 
over  a  zone  of  half  to  one  centimetre ;  the  necrotic  patch 
becomes  detached,  and  the  ulcer  heals  up  and  closes  without 
any  swelling  of  the  corresponding  glands.  The  same  process 
takes  place  when,  instead  of  living  bacilli,  bacilli  killed  by 
boiling  are  injected  the  second  time. 

The  tubercle  bacilli  therefore  act  in  a  different  fashion  in 
the  healthy  organism  from  the  organism  already  tuberculous. 
Koch  observed  that  a  large  dose  of  bacilli  killed  the 
tuberculous  guinea-pig,  whereas  a  very  small  dose  produced 
an  improvement  in  their  condition  and  healed  up  the  initial 
ulceration.  He  saw  in  this  a  principle  of  treatment.  Since 
the  bacilli  are  not  readily  absorbed,  he  replaced  them  by  an 
extract  from  cultures  ;  this  was  the  original  tuberculin. 

This  substance  is  practically  harmless  to  non-tuberculous 
animals,  but  is  fatal  to  tuberculous  animals  in  a  very  small 
dose.  In  absolutely  minimal  doses,  repeatedly  administered,  it 
exerts  a  curative  effect  on  certain  tuberculous  lesions.  Accord- 
ing to  the  size  of  the  dose,  it  can  act  as  a  poison  or  as  a  remedy. 
These  are  its  fundamental  properties. 

Local  and  General  Reactions. — Koch  ascribed  the 
curative  effect  to  the  necrotic  action  which  he  had  noticed 
after  reinoculation  of  bacilli  in  tuberculous  guinea-pigs. 
Tuberculin  he  said  in  1890,  does  not  kill  the  bacilli,  it  kills 
the  living  tuberculous  tissues ;  it  does  not  even  act  on  tissues 
already  dead,  such  as  the  caseous  masses.  It  acts  on  cells  in 
the  same  way  as  the  bacillus  tuberculosis  itself,  but  the  soluble 
product  has  a  much  more  extensive  radius  of  activity  than  the 
bacillus. 

In  what  way  does  this  necrotic  action  become  therapeutic  ? 
Because  in  a  necrotic  tissue  the  bacillus  is  badly  nourished 
and  grows  feebly ;  the  dead  tissue  becomes  a  sort  of  slough 
or  sequestrum  which  the  body  strives  to  get  rid  of.  The 
action  of  tuberculin  is,  in  a  sense,  surgical.  It  might  be 
hoped  that  every  part  affected  by  it  might  be  thrown  off,  and 
this  is  sometimes  possible  in  tuberculosis  of  the  skin  and  of 
the  lungs.  In  many  cases,  however,  it  is  impossible,  and  it 


TUBERCULIN   AND   MALLEIN  175 

has  always  been  feared  that  tuberculin  might  cause  a  necrosis 
of  the  tuberculous  tissue  without  completely  killing  the  bacilli, 
and  might  thus  set  them  free  and  inoculate  them  on  a  tissue 
till  then  unaffected. 

When  the  tuberculin  action  does  not  quite  reach  the  degree 
of  necrosis,  it  merely  produces  around  the  tuberculous  focus 
an  active  inflammatory  reaction  with  an  afflux  of  leucocytes, 
which  may  build  up  a  fibrous,  cicatricial  tissue  in  place  of  the 
tuberculous  ulceration. 

Tuberculin  does  not  produce  only  local  reactions;  it  pro- 
vokes a  general  reaction  of  the  body,  the  most  obvious  sign 
of  which  is  fever.  In  large  doses  the  reaction  occurs  even  in 
a  healthy  individual  according  to  R.  Koch,  who  observed  this 
in  himself ;  but  it  is  very  probable  that  he  had  at  that  time 
some  tuberculosis,  and  it  has  been  maintained  that  tuberculin 
is  entirely  inactive  in  subjects  who  have  never  been  attacked 
by  the  bacillus. 

Three  or  four  hours  after  the  inoculation  of  ^  c.c.,  Koch 
observed  "twinges  of  pain  in  the  limbs,  a  feeling  of 
fatigue,  and  a  tendency  to  cough.  The  symptoms  became 
more  pronounced,  and  about  the  fifth  hour  he  had  a  violent 
rigorwhich  lasted  awhole  hour  with  general  uneasiness,  vomiting, 
and  fever  (103° -3  F.).  The  symptoms  began  to  settle  about 
the  twelfth  hour,  and  on  the  following  day  the  temperature  was 
normal ;  a  heaviness  of  the  limbs  and  stiffness  were  perceptible 
for  several  days  after.  The  point  of  inoculation  remained  red 
and  painful  for  a  considerable  time." 

In  the  treatment  of  a  tuberculous  patient  with  tuberculin, 
doses  are  employed  which  do  not  produce  these  violent 
symptoms.  As  far  as  possible  no  symptom,  not  even  fever, 
ought  to  occur.  A  well-conducted  treatment  produces  an 
improvement  in  many  patients  ;  this  fact  is  certain,  but  the 
mechanism  of  these  cures  is  not  yet  well  understood.  The 
indications  and  contra-indications  for  treatment  are  complex, 
and  cannot  be  settled  except  by  a  physician  with  a  large 
experience  of  tuberculosis  and  tuberculin  treatment. 

The  febrile  reaction  which  follows  the  inoculation  of  a  dose 


176  MICROBES   AND  TOXINS 

too  small  to  be  dangerous  is  employed  in  the  diagnosis  of 
tuberculosis,  both  among  animals  and  in  man. 

Tuberculin  is  very  different  from  other  toxins.  It  bears 
much  less  resemblance  than  they  to  the  ferments.  In  the 
liquid  condition  it  stands  a  temperature  of  120°  to  150°  C. 
In  the  solid  state,  heated  dry  in  sealed  tubes,  it  stands  250°  C. 
"  If  it  is  a  substance  derived  from  albumins,"  said  Koch,  "  it 
cannot  be  a  toxalbumin  in  view  of  this  resistance  to  heat  and 
of  its  dialysing  properties/'  It  may  be  exposed  to  sunlight  for 
months  without  losing  its  activity.  Heating  with  acids  (for 
example,  -gVtn  hydrochloric)  and  with  alkalies  simply  weakens 
it  without  destroying  it.  Perhaps  it  is  not  a  simple  poison  : 
in  the  condition  in  which  we  get  it,  it  has  no  more  claim  to  be 
pure  than  our  diphtheria  and  tetanus  toxins.  Maragliano 
thinks  that  it  contains,  besides  the  poison  which  causes  the 
fever  and  is  not  a  toxalbumin,  a  poison  which,  on  the  contrary, 
lowers  the  temperature  of  the  body,  is  destroyed  by  heating  to 
100°  C.,  and  is  possibly  a  true  toxalbumin. 

Tuberculin  without  tubercle  bacilli  does  not  reproduce 
tuberculosis.  To  provoke  suppuration,  caseation,  and  the 
typical  lesion,  the  tubercle,  living  or  dead  tubercle  bacilli  are 
necessary :  fluid  tuberculin  does  not  even  produce  the  lesions 
which  dead  bacilli  can  give  rise  to  :  it  has  nothing  in  common 
with  them  but  its  destructive  action  on  the  cells  and  its  power 
of  raising  temperature. 

It  is  eminently  specific,  not  producing  any  definite  effects 
except  in  tuberculous  individuals.  In  this  it  differs  entirely 
from  the  other  bacterial  poisons.  It  only  acts  on  a  prepared 
soil,  a  soil  prepared  by  the  bacillus  tuberculosis  itself,  i.e.,  by 
an  agent  to  which  it  is  closely  related  both  by  origin  and 
constitution.  This  specificity,  though  very  marked,  is  not 
absolute.  Tuberculin  acts  similarly,  less,  it  is  true,  than  in 
tuberculous  individuals,  but  more  than  in  healthy  subjects,  on 
patients  affected  by  lesions  resembling  anatomically  the 
tubercle,  e.g.,  glanders  and  actinomycotic  nodules.  This 
extension  of  its  field  of  action  is  perhaps  due  to  the  close 
biological  relationship  between  the  tubercle  bacillus  and  the 


TUBERCULIN   AND  MALLEIN  177 

micro-organisms  of  these  diseases,  perhaps  to  the  existence  of 
the  same  anatomical  type  of  lesion,  the  tubercle,  or  perhaps 
to  similar  inflammatory  and  phagocytic  reactions. 

Antituberculin  ? — Does  tuberculin  in  animals,  tuber- 
culous or  not,  treated  and  habituated  to  its  effects,  give  rise  to 
the  production  of  an  antituberculin  comparable  to  antitoxins 
or  even  anti-endotoxins ?  No;  an  antituberculous  serum 
comparable  to  antidiphtheria  or  even  antiplague  sera  does  not 
exist,  in  spite  of  all  efforts  to  discover  it.  Tuberculin  in 
tuberculous  subjects  excites  the  production  of  certain  reaction- 
or  anti-bodies,  but  no  true  antitoxin.  No  treated  individual 
has  ever  furnished  a  serum  capable  of  neutralizing  tuberculin 
either  in  vitro  or  in  vivo,  but  serum  of  this  kind  can  produce 
precipitates  and  clumps  in  a  suspension  of  bacilli ;  this  action 
is,  however,  inconstant  and  of  little  value  in  medicine. 

Wasserman  and  Bruck  have  given  the  name  of  antituberculin 
to  the  reaction  products  which  fix  themselves  on  tuberculin  as 
anti-bodies  do  on  antigens  (v.  Chap.  X.).  The  interpretation 
of  these  experiments  is  a  question  of  some  delicacy,  and  we 
shall  see  how  doubtful  are  the  relations  between  the  presence 
of  antibodies  and  the  existence  of  an  immunity,  e.g.,  a 
resistance  in  the  case  of  tuberculous  individuals. 

Habituation  to  Tuberculin. — By  careful,  repeated 
injections  the  tuberculous  subject  may  be  made  to  protect 
himself  against  the  fatal  action  of  tuberculin,  and  to  enjoy  a 
general  improvement  in  his  condition,  without  stopping  the 
progress  of  his  tuberculosis.  Guinea-pigs  may  be  made  to 
support  fifty  lethal  doses,  yet  their  lesions  progress  in  the 
ordinary  way — or  even  more  quickly  than  usual.  It  is  not 
yet  well  known  what  are  the  relations  between  the  physiological 
action  of  tuberculin  and  the  progress  of  a  chronic  tuberculosis. 

This  habituation  has  been  called  immunity  to  tuberculin.  It 
is  not,  however,  an  immunity  or  even  a  resistance  to  tuberculosis. 
It  does  not  appear  on  repeated  injections  of  equal  small  doses 
of  tuberculin  ;  in  this  case,  the  febrile  reaction,  which  was 
absent  at  first,  finally  appears  and  becomes  severe.  The  fever 
remains  absent  when  one  proceeds  by  increasing  doses  (always 


178  MICROBES   AND  TOXINS 

to  be  done  with  caution).  With  small  equal  repeated  doses, 
tuberculin  behaves  like  a  poison  towards  which  the  tuberculous 
patient  becomes  more  and  more  sensitive. 

Cutaneous  Reactions. — A  drop  of  very  dilute  tuberculin, 
applied  by  means  a  prick  or  very  superficial  scarification  on  the 
non-tuberculous  skin  of  a  tuberculous  subject,  excites  at  the 
point  a  reaction  which  may  extend  to  the.  vessels  and  lympathic 
glands  of  the  neighbourhood — V.  Pirquet's  experiment ;  this 
represents  a  new  diagnostic  procedure,  the  cuti-reaction.  It  has 
been  modified  and  perfected  by  dropping  the  tuberculin  between 
the  eyelids  (conjunctiva  or  oculo-reaction  of  Wolff-Eisner  and  of 
Calmette)  or  by  inoculating  it  with  a  very  fine  needle  in  the 
depths  of  the  skin  itself  (the  intra-dermo-reaction  of  Mantoux). 

It  is  a  reaction  of  extreme  interest,  for  it  occurs  at  a  non- 
tuberculous  point,  i.e.,  a  point  containing  no  bacilli,  in  a 
tuberculous  individual ;  and  can  only  be  explained  by  suppos- 
ing that  the  whole  body  has  become  impregnated  with  sub- 
stances formed  under  the  influence  of  the  tubercle  bacillus. 


ANIMAL  TOXINS — THE  VENOMS  1 

In  the  struggle  for  existence,  certain  animal  species  have 
acquired,  as  their  means  of  attack  and  defence,  organs  which 
secrete  and  inoculate  toxic  substances.  These  animal  toxins 
are  the  venoms,  and  such  venoms  are  known  at  almost  every 
level  in  the  animal  scale. 

Venoms  in  the  Animal  World. — Among  the  Ccelen- 
terata  the  Actinians  produce  certain  poisons  which  can  be 
extracted  from  their  tentacles,  thalassine  and  congestine^  well 
known  as  having  formed  the  subject  of  Ch.  Richet's  experi- 
ments on  anaphylaxis.  These  poisons  are  perhaps  the  cause  of 
the  disease  of  sponge-fishers,  who  dive  quite  naked  without  a 
diving-suit ;  the  disease  consists  in  burning  of  the  skin  and 
swelling,  with  gangrene  and  violent  fever. 

The  pedicles  of  the  Sea-Urchins  (Echinoderms)  contain  a 

1   Vide  papers  of  Noguchi  and  Calmette. 


TUBERCULIN  AND   MALLEIN  179 

poison  which  stands  boiling.  This  poison,  in  nature,  is 
dangerous  to  crabs  and  fishes ;  in  the  laboratory  it  is  found 
to  kill  the  rabbit. 

Among  the  Arthropoda,  the  Spiders  and  Scorpions  (Arach- 
nidae)  secrete  active  poisons.  The  excretory  tubes  of  the 
poison  glands  of  venomous  spiders  open  at  the  point  of  the 
two  appendices  which  are  furnished  with  claws  at  the  end  and 
situated  on  each  side  of  the  mouth.  The  poison  kills  the 
small  animals  on  which  the  spiders  feed,  and  causes  in  man 
pain  and  contracture  at  the  bitten  point — a  sort  of  miniature 
tetanus.  The  poison  of  certain  spiders  contains  a  haemolysin^ 
i.e.)  it  lakes  blood,  making  the  haemoglobin  of  the  corpuscles 
diffuse  into  the  surrounding  liquid  (arachnolysin).  The  bite 
of  the  Tarantula  (Lycosa  tarentula)  is  only  dangerous  for  the 
small  animals  on  which  they  feed,  and  is  quite  harmless  to 
man.  According  to  Brehm,  all  the  stories  of  the  effects  on 
man  of  the  Tarantula  bite  are  nothing  but  fables  and 
fantasies. 

The  venom  of  the  Scorpion  (Scorpio  occitanus)  of  the  South 
of  France  can  kill  a  guinea-pig  in  a  dose  of  half  a  milligram  of 
dry  extract ;  for  a  rabbit  one  milligram.  The  scorpion  is  the 
subject  of  a  legend  which  says  that  when  it  is  enclosed  by  a 
circle  of  fire  it  commits  suicide  with  its  own  poison.  Now 
the  scorpion  is  in  reality  immune  to  scorpion  venom,  towards 
which  its  serum  acts  like  an  antitoxin  (Metchnikoff). 

Among  the  Myriapoda  the  Centipedes,  and  among  insects  the 
Hymenoptera,  secrete  venoms.  The  poison  extracted  from  two 
bees  (by  grinding  up  the  terminal  part  of  their  bodies  in  i  c.c. 
of  water)  is  sufficient  to  kill  by  asphyxia  a  mouse  or  a 
sparrow.  It  also  is  a  haemolytic  poison. 

There  are  many  poisonous  fishes.  As  a  rule  their  poison 
glands  are  found  at  the  base  of  the  dorsal  or  caudal  fins,  or 
under  the  spine  of  the  gill-flap.  These  venoms  all  resemble 
more  or  less  that  of  the  weever-fish,  which  has  been  most 
studied.  Locally  it  causes  pain  and  swelling,  with  fever  and 
vomiting.  At  the  time  of  spawning  the  poison  is  more 
abundant  and  more  active.  The  tropical  tetrodons  are  most 

N    2 


180  MICROBES  AND  TOXINS 

venomous  at  the  time  of  greatest  activity  of  their  reproductive 
glands. 

The  Toad  (Batrachia)  manufactures  poison  in  its  parotid 
gland  and  the  glands  of  the  skin,  but  it  has  no  other  way  of 
secreting  it  than  by  contracting  its  skin  and  covering  itself  with 
a  viscous,  nauseating  slime,  which  is  rather  poisonous  when 
injected  into  small  animals  such  as  mice.  Phisalix  and 
Bertrand  have  extracted  two  poisons  from  the  toad,  '  bufotaline ' 
and  '  bufotenincj  a  poison  of  the  nervous  system.  At  spawning 
time  the  cutaneous  glands  of  the  male  toad  are  full  of  venom, 
whereas  those  of  the  female  are  empty;  but  the  poison 
accumulates  in  her  eggs,  from  which  it  may  be  extracted  by 
means  of  chloroform. 

The  Salamanders  possess  on  their  sides  and  tail  poison 
glands,  and  it  is  to  the  fluid  which  these  secrete  that  they  owe 
their  fame  as  animals  capable  of  living  in  fire  and  even 
of  extinguishing  it — pure  legend,  of  course.  Their  secretion 
permits,  at  most,  of  their  surviving  a  few  seconds. 

One  venomous  animal  exists  among  the  mammals,  the 
Ornithorhynchus.  Its  poison  gland  is  situated  on  its  thigh, 
and  the  secretion  escapes  by  a  spur  or  claw  on  the  hind  feet. 
The  poison  resembles  that  of  the  snake  Lachesis^  but  is  much 
more  feeble. 

Medical  men  have  been  especially  attracted  to  the  study 
of  snake-venoms.  The  vipers  of  our  own  country  have  few 
victims,  but  in  India  the  cobra  kills  as  many  as  an  epidemic 
disease.  In  1889,  in  India,  22,480  human  beings  and  3,793 
domestic  animals  died  of  snake-bite.  Of  those  bitten  25  to  30 
per  cent,  die  within  *ten  or  twelve  hours.  The  importance 
is  obvious  of  the  researches  which  led  to  the  antivenom 
serotherapy  (Calmette). 

As  they  issue  from  the  glands  the  venoms  resemble  a  thick, 
oily  saliva  more  or  less  yellow.  Their  physical  properties  vary 
a  good  deal  with  the  genus.  The  venoms  of  the  Viperidae 
do  not  dialyse  through  a  membrane  and  are  destroyed  entirely 
at  75°  to  80°  C.  (Lachesis  even  at  65°).  Those  of  the 
Colubridae  pass  slowly  through  vegetable  membranes,  with 


TUBERCULIN   AND  MALLEIN  181 

greater  difficulty  through  animal  parchment :  they  resist  heating 
at  100°  C.  and  are  only  completely  destroyed  at  115-120°. 

From  the  chemical  point  of  view  both  consist  of  proto- 
and  deutero-albumoses :  the  albumins  they  contain  are  not 
toxic.  S.  Faust  has  extracted  from  cobra  venom  an  "  ophio- 
toxin"  which  contains  neither  nitrogen  nor  sulphur  nor 
phosphorus. 

By  using  dried  venom,  which  can  be  accurately  dosed  by 
redissolving  in  a  known  volume,  it  has  been  possible,  as  with  the 
vegetable  toxins,  to  determine  the  minimal  lethal  dose  per 
kilogram  in  different  species  of  animal.  As  with  tetanus  toxin 
the  size  of  the  animal  bears  no  relation  to  its  sensitiveness. 
With  one  gram  of  dry  cobra-venom  it  is  possible  to  kill  1,250 
kilos,  weight  of  dog,  2,000  k.  of  rabbit,  2,500  k.  of  guinea-pig, 
1,430  k.  of  rat,  8,333  k.  of  mouse,  20,000  k.  of  horse,  and 
10,000  of  man,  i.e.,  165  adults  of  average  weight.  The  horse 
is  thus  the  most  sensitive  of  all  these  animals. 

The  toxicity  of  the  venom  is  very  variable  :  it  is  more 
active  (and  doubtless  less  abundant)  after  the  casting  of  the 
skin  and  after  a  prolonged  fast. 

Physiological  Action  of  Snake-venoms.  —  The 
physiological  action  of  snake-venoms  is  complex.  They  act 
on  the  cells  of  the  organs,  on  the  liver,  the  kidney,  and  the 
spleen :  on  the  endothelial  cells  which  line  the  interior  of  the 
blood  vessels  (especially  the  rattlesnake  poisons) :  on  the  nervous 
system  (according  to  Rogers  the  venoms  of  the  Viperidae 
paralyse  the  vasomotor  centres,  those  of  the  Colubridae  the 
respiratory  centres) :  on  the  blood,  one  of  the  oldest  known 
effects  and  one  much  studied  recently  because  the  solution  of 
the  red  corpuscles  or  haemolysis  is  a  phenomenon  very  obvious 
and  easy  to  study  in  vitro.  The  condition  of  the  blood  at 
autopsy  varies  according  to  the  dose  of  the  poison  and  the 
time;  this  is  why  the  same  poison  is  called  coagulant  or' 
anticoagulant  by  different  workers. 

Snake-venoms  have  the  properties  of  digestive  ferments. 
They  can  dissolve  coagulated  blood  and  can  destroy  the  cells 
of  the  vessel  coats  and  even  of  the  muscles.  Cobra-venom 


182  MICROBES   AND  TOXINS 

digests  albumins  but  without  reaching  the  peptone  stage.  The 
digestive  properties  are  destroyed  by  heating  to  70°  C. 

Pancreatic  juice,  as  is  well  known,  when  perfectly  pure 
cannot  alone  digest  albumin ;  it  has  to  be  "  activated "  by  a 
"kinase  "  ferment,  secreted  either  by  the  intestinal  mucosa  or 
by  the  leucocytes.  Now  snake-venoms  can  take  the  place  of 
this  kinase  and  activate  a  pure  inactive  pancreatic  juice.  This 
is  all  the  more  curious  since  the  pancreatic  juice  when 
activated  digests  and  destroys  venoms,  which,  as  a  rule,  have 
in  consequence  no  action  when  taken  by  the  mouth.  The 
venom  secretion  is  thus  for  the  snake  itself  a  normal 
physiological  secretion  of  great  use  in  the  digestion  of  the 
huge  meals  for  which  snakes  are  famous.  There  is  further 
nothing  surprising  in  the  fact  that  non-venomous  snakes,  i.e., 
those  not  provided  with  poison-fangs,  still  possess  glands 
capable  of  secreting  venom ;  it  is  simply  used  in  the  digestion 
of  their  food. 

Venoms  thus,  like  toxins,  resemble  ferments — with  the  same 
reservations.  They  are  very  closely  allied  to  toxins.  Their 
action,  like  that  of  toxins,  is  not  simple;  just  as  in  tetanus 
toxin  several  different  substances  or  functions  can  be  dis- 
tinguished, a  nerve-cell  poison  and  a  poison  of  the  red  blood 
corpuscles,  so  several  physiological  activities  can  be  dis- 
tinguished in  the  same  venom.  But  venoms  act  without  any 
incubation  period,  or  at  any  rate  with  a  very  short  one.  It  is 
only  the  time  elapsing  between  inoculation  and  death  that 
varies  with  the  dose,  and  that  within  rather  narrow  limits. 

Like  the  toxins,  the  venoms  are  destroyed  at  relatively  low 
temperatures  (resistance  up  to  100°  to  110°  C.  does  not  interfere 
with  the  analogy).  They  act  in  minute  dose  and  deteriorate 
or  are  destroyed  by  light,  by  photodynamic  substances,  by 
iodine  perchloride,  and  alkaline  hypochlorites. 

Finally,  and  most  important,  the  venoms  give  rise  to 
antivenoms,  as  toxins  do  to  antitoxins.  It  is  practically  only  with 
the  help  of  the  antivenoms  indeed  that  the  specificity  of  the 
venoms  can  be  definitely  demonstrated. 

Venom     Haemolysis.— The     venom      are    haemolytic 


TUBERCULIN   AND   MALLEIN  183 

poisons,  and  since  haemolysis  is  a  phenomenon  much  more 
convenient  to  follow  experimentally  than  paralyses  or  nervous 
symptoms,  they  have  been  much  studied,  and  from  certain 
points  of  view  are  better  known  than  the  microbial  toxins. 
The  study  of  the  toxins  has  profited  by  that  of  the  venoms. 
Thanks  to  some  beautiful  experimental  results,  there  is  reason 
to  believe  that,  with  the  help  of  the  venoms,  the  study  of 
toxins  in  general  may  make  the  advance  so  much  desired  by 
science,  and  from  a  physiological  subject,  studied  only  in  the 
living  animal,  become  a  part  of  chemistry  with  its  definite 
reactions. 

Some  definitions  and  examples  must  first  be  given. 

When  a  rabbit  is  repeatedly  inoculated  with,  for  example, 
defibrinated  sheep's  blood,  the  rabbit's  serum  acquires  the 
property  of  dissolving  the  red  corpuscles  of  the  sheep  :  these 
latter  suspended  in  physiological  saline  solution  in  a  test-tube 
with  a  little  of  this  rabbit's  serum,  instead  of  settling  intact 
and  leaving  a  colourless  supernatant  fluid,  break  up,  liberate 
their  haemoglobin,  and  colour  the  fluid  red.  The  rabbit's 
serum  has  become  haemolytic  for  the  red  corpuscles  of  the 
sheep. 

Bordet  has  shown  that  haemolysis  depends  on  the  operation 
of  two  substances,  or  rather  of  two  functions,  of  which  we 
shall  have  much  to  say  in  connection  with  immunity  :  one  is 
the  alexine  or  complement  of  normal  serum  (destroyed  by 
heating  to  56°  C.  for  one  hour)  ;  the  other  is  the  sensibilisatrice 
or  immune-body  of  the  serum  of  immunised  animals,  such 
as  the  rabbit  above  mentioned  (it  stands  heating  to  68°  or 

70°  c.). 

The  latter  owes  its  name  to  the  fact  that  it  prepares  or 
renders  sensitive  the  red  corpuscles  towards  the  action  of  the 
complement.  The  complement  completes  the  action  of  the 
"sensibilisatrice,"  hence  its  name.  If  we  take  the  blood 
corpuscles  of  a  goat  and  add  a  little  cobra-venom,  haemolysis 
occurs.  But  if  the  blood  corpuscles  are  first  carefully  washed 
with  physiological  saline  so  as  to  be  entirely  freed  from  traces 
of  blood  serum  which  might  adhere  to  them,  no  haemolysis 


184  MICROBES   AND   TOXINS 

occurs  when  the  venom  is  added.  But  if  now  to  this  mixture 
of  carefully  washed  goats'  corpuscles  and  venom  a  little  normal 
blood  serum  is  added,  haemolysis  proceeds  at  once.  It  would 
seem  from  these  facts  that  the  venom  acts  like  a  sensibilisatrice 
or  immune-body,  the  normal  serum  providing  the  alexine  or 
complement. 

But  there  are  other  facts  which  forbid  this  interpretation. 
Normal  serum  activates  the  venom,  it  is  true,  but  even  when 
heated  to  65°  C.  or  higher  it  still  activates :  there  exist  even 
normal  sera  which  cannot  activate  venom  haemolysis  until  they 
have  been  heated  at  100°  C.  It  is  inconceivable  that  it  is  the 
complement  which  is  the  active  agent  since  complement  is 
destroyed  at  56°  C. 

Further,  washed  corpuscles  of  certain  animals  are  laked  by 
the  venom  without  any  addition  of  fresh  serum  (corpuscles  of 
dog,  rat,  guinea-pig,  and  man).  Again,  in  the  case  of  the 
washed  goats'  corpuscles,  normal  serum  is  not  the  only 
substance  which  can  activate  :  laked  red  corpuscles,  e.g.,  of  the 
guinea-pig,  can  take  its  place  quite  well.  Finally,  in  this  last 
example  it  is  not  the  fluid  which  acts,  but  the  stromata  or 
bodies  of  the  corpuscles  which  have  shed  their  haemoglobin, 
and  these  still  possess  their  activity  after  heating  to  100°  C. 
It  is  thus  impossible  to  attribute  the  activating  action  to  the 
complement  or  even  to  the  serum  as  a  fluid.  The  active 
substance  is  not  the  complement  nor  is  it  an  albumin,  for 
albumin  coagulates  below  100°  C.  It  is  not  a  ferment,  for  a 
ferment  heated  in  solution  above  100°  C.  is  no  longer  active. 
It  is  a  definite  chemical  substance  present  in  the  serum  and 
in  the  stroma  of  the  blood  corpuscles,  namely  lecithin. 
Lecithin  is  a  well-defined  chemical  body,  unlike  albumin,  for 
which  we  are  unable  to  write  a  formula,  still  more  unlike  the 
complement  and  the  immune  body  which,  like  the  ferments,  are 
known  as  activities  or  properties,  not  as  substances.  In  venom 
haemolysis,  therefore,  our  knowledge  is  more  complete  and 
clear  than  in  the  haemolysis  of  haemolytic  sera. 

Lecithids. — When  blood  of  any  species  is  easily  laked 
by  venom  without  addition  of  serum  (e.g.,  in  man,  the  rat, 


TUBERCULIN   AND   MALLEIN  185 

and  the  guinea-pig)  it  means  that  the  lecithin  of  the  corpuscles 
readily  detaches  itself  and  unites  with  the  venom.  When  the 
blood  is  laked  only  when  serum  is  furnished  in  addition,  it 
means  that  the  lecithin  of  the  globules  themselves  is  firmly 
bound  and  difficult  to  set  free.  In  certain  cases  heating  to 
65°  C.  or  even  higher  is  necessary  to  liberate  lecithin  from  its 
combination  with  haemoglobin.  The  experiments  of  this 
kind  have  many  complications  of  detail,  since  the  three  factors 
coming  into  play— the  blood  corpuscles,  the  lecithin,  and  the 
venom — are  subject  to  many  variations.  A  step  in  advance 
has  been  made  in  what  may  be  called  the  mechanical  or 
purely  chemical  explanation  of  haemolysis  by  the  discovery 
that  lecithin  forms  with  venom  a  combination  of  a  chemical 
character  in  which  neither  lecithin  nor  venom  can  be 
recognized.  Kyes  has  named  this  combination  or  "  couple  " 
lecithid)  or  since  his  experiments  were  on  cobra-venom,  cobra- 
lecithid. 

In  its  physical  properties  (solubility  in  water,  alcohol,  ether, 
chloroform,  and  acetone)  the  lecithid  differs  from  lecithin  as 
much  as  from  venom.  It  can  be  isolated  in  a  crystalline  form 
and  redissolved  in  water.  It  acts  on  the  blood  of  all  animal 
species,  and  that  without  any  incubation  period.  The  delay 
observed  in  the  action  of  venoms  is  not  a  period  of  incubation, 
but  merely  represents  the  time  necessary  for  the  formation  of 
the  lecithid  compound.  If  the  ready-made  lecithin  is  added 
to  the  blood  the  haemolysis  is  more  rapid  than  on  the  addition 
of  the  two  elements  separately. 

Strictly  speaking,  however,  the  venoms  do  not  act  without 
incubation  :  the  time  taken  by  lecithid  formation  represents 
the  minimum  incubation  period.  It  is  quite  possible  that  in 
the  action  of  microbial  toxins  there  may  occur  a  slow  forma- 
tion (i.e.,  with  a  longer  incubation  period)  of  compounds 
analogous  to  lecithids. 

Cholesterin  behaves  as  an  antagonist  to  lecithin  ;  it  has  no 
effect  on  complement,  but  prevents  the  combined  action  of 
the  lecithid  by  affecting  the  lecithin ;  it  thus  forms  a  sort  of 
antihczmolysin  or  antitoxin  the  composition  of  which  is  definitely 


186  MICROBES   AND  TOXINS 

known.  It  is  one  of  the  dreams  of  biological  chemistry  to 
discover  an  equivalent  to  cholesterin  for  the  other  toxins. 

Still  more  interesting  is  the  fact  that  the  peculiar  anaemia 
produced  by  injecting  cobra-lecithid  into  an  animal  can  be 
prevented  by  giving  cholesterin.  It  would  seem  that  cholesterin 
acts  towards  lecithin  and  lecithid,  not  only  as  an  antagonist, 
but  as  a  remedy. 

There  are,  of  course,  facts  which  prevent  us  from  regarding 
as  entirely  similar  the  cobra-hsemolysin,  on  the  one  hand,  and 
the  solvent  properties  or  haemolysins  of  normal  and  immune 
sera  (sera  derived  from  animals  prepared  by  the  injection  of 
blood  corpuscles)  on  the  other.  These,  however,  do  not 
invalidate  Kyes's  conclusions  in  his  experiments  on  the 
lecithids.  The  study  of  snake-venoms  has  shown  a  participa- 
tion of  chemically-defined  substances  in  phenomena  hitherto 
only  known  from  the  biological  point  of  view.  It  would  be 
of  immense  interest,  both  theoretical  and  practical,  to  discover 
an  analogous  mechanism  in  the  effects  of  toxins. 

Certain  animals  enjoy  a  natural  immunity  towards  snake- 
venom,  which,  however,  is  never  absolute.  It  is  possible  that 
these  animals  have  received  from  generation  to  generation 
small  doses  of  the  venom  as  the  result  of  being  bitten,  and 
that  they  have  in  consequence  elaborated  an  antivenom. 
This  forms  the  starting-point  of  artificial  immunisation  and 
antivenom  serum-therapy.  It  is  known  that  the  blood  of 
certain  animals  possesses  normally  a  weak  antitoxic  action 
against  diphtheria  and  tetanus  toxins ;  these  animals  similarly 
must  have  harboured  tetanus  and  diphtheria  bacilli. 

The  animals  possessing  the  most  remarkable  resistance 
towards  snake-venoms  are  the  hedgehog  and  the  mongoose. 
To  see  the  dramatic  combats  which  take  place  when  the 
mongoose  tackles  the  cobra,  one  has  only  to  read  Kipling's 
marvellous  tale  of  the  war  between  "  Rikki-Tikki "  and  "  Nag  " 
in  \hejungle  Book, 


CHAPTER  X 

IMMUNITY 
PHAGOCYTOSIS  AND  HUMORAL  IMMUNITY 

Early  ideas  of    Pasteur    on    immunity — Opposition    to    the    phagocytic 

doctrine — Cellular  and  humoral  immunity. 
Antigens    and   Antibodies — Complement — The   two  substances  :  Borders 

experiments. 
Phagocytosis  a  fact    capable    of   direct    observation — Ferments    of   the 

leucocytes — Analogies  with  the  digestive  ferments. 
Pfeiffer's    phenomenon— Opsonins    and    bacteriotropins — Antibodies   not 

an  exact  measure  of  the  immunity. 

IT  is  not  necessary  to  have  studied  medicine  or  science 
in  order  to  ask  oneself  what  is  this  immunity  which  appears  in 
infectious  disease.  Certain  bacteria  are  pathogenic  for  certain 
animal  species  and  not  for  others ;  the  guinea-pig,  for  example, 
does  not  take  fowl-cholera  and  the  fowl  does  not  take  anthrax. 
Among  people  living  in  one  family  under  the  same  conditions 
and  among  soldiers  in  the  same  barracks  living  under  the 
same  rules  we  see  disease  attacking  some  while  others  remain 
free.  Finally  it  is  a  popular  conviction  that  anthrax  does 
not  occur  twice  and  that,  as  a  rule,  once  an  individual  has 
had  measles  or  small-pox  he  never  takes  it  again:  these 
are  everyday  examples  of  acquired  immunity. 

"  Immunity  against  infectious  diseases  ought  to  be 
understood  to  mean  the  sum  total  of  all  the  phenomena 
to  which  is  due  the  resistance  of  an  animal  body  to  the 
microbes  which  produce  these  diseases "  (Metchnikoff). 
Immunity  may  be  innate  or  acquired.  Natural  acquired 

187 


188  MICROBES   AND  TOXINS 

immunity  appears  when  there  is  spontaneous  recovery  from  an 
infectious  disease.  Immunity  the  result  of  human  interference 
(vaccinations,  serotherapy)  is  artificial  acquired  immunity. 

After  his  work  in  collaboration  with  Chamberland  and  Roux 
which  established  the  attenuation  of  viruses  and  the  value  of 
preventive  inoculations,  Pasteur,  being  a  chemist,  conceived 
immunity  as  a  chemical  process.  He  considered  that  the 
reason  why  the  bacillus  of  fowl-cholera  fails  to  grow  in 
the  fowl  vaccinated  against  this  disease  was  that  the  body 
of  such  a  fowl  no  longer  contained  the  necessary  food-stuffs 
for  the  development  of  the  microbe.  The  muscle  which  has 
been  severely  affected  by  disease  has  become,  even  after 
complete  recovery,  in  some  way  incapable  of  supporting  the 
life  of  the  microbe,  as  if  this  latter  during  its  previous  growth 
had  made  to  disappear  from  the  muscle  some  principle  which 
life  is  incapable  of  renewing  and  the  absence  of  which  prevents 
the  development  of  the  micro-organism.1 

He  filtered  a  culture  of  the  fowl-cholera  bacillus  and  found 
that  a  re-inoculation  of  the  bacterium  in  the  fluid  thus  freed 
from  the  first  germs  always  failed :  when  fresh  nutritive 
substances  were  added  to  the  filtrate,  growth  took  place.  It  was 
not,  therefore,  the  presence  of  some  excretion,  but  the  absence 
of  some  nutritive  substance  which  explained  "  the  immunity  of 
a  culture  filtrate,  or  of  the  fowl  considered  as  a  natural  culture 
medium." 

In  natural  innate  immunity  also  he  refused  to  recognise  the 
presence  of  an  inhibitory  substance,  basing  his  faith  on  the 
celebrated  experiment  of  the  fowl  refractory  to  anthrax  but 
rendered  susceptible  by  chilling,  and  appealing  to  the  "con- 
stitution "  or  to  a  "  vital  resistance,"  by  which  he  conceived  a 
struggle  between  the  parasites  and  the  body-cells  for  the 
oxygen  and  food  materials  available.  But  when  it  was  found 
that  bacteria  could  grow  perfectly  in  the  blood  of  animals 
possessing  a  complete  immunity,  Pasteur's  early  conception 
could  no  longer  be  maintained  in  its  primitive  simple  form. 

Even  to-day  immunity  has  still  to  be  defined  as  a  complex 
1  C.  R.  Acad,  de$  Sciences,  1880,  p.  247. 


IMMUNITY  189 

of  biological  phenomena,  for  in  spite  of  the  hope  of  men  of 
science  some  day  to  get  beyond  "  vitalistic "  explanations,  no 
explanation  in  chemical  terms  can  yet  be  given.  Immunity  is 
a  function  of  the  cells.  Immunity  means  phagocytosis.  Further 
research  may  fathom  the  nature  of  this  activity  and  give  a 
chemical  explanation  as  in  the  case  of  peptic  or  pancreatic 
digestion,  but  the  cellular  activity  is  indisputable  and  is  not  a 
theory  but  a  collection  of  facts,  a  doctrine  in  the  true  sense. 

The  principles  of  this  doctrine  of  phagocytic  immunity  have 
already  been  indicated  in  the  chapter  on  inflammation.  It  is 
necessary  to  read  in  MetchnikofFs  book  his  "  historical  review 
of  our  knowledge  on  immunity  "  (Chap.  XVI)  to  comprehend 
how  much  his  doctrine  has  developed. 

From  the  historical  point  of  view  it  had  to  oppose  the  ruling 
conceptions,  not  only  in  medicine,  but  in  pathological  anatomy 
and  in  physiology.  The  few  observers  who  had  seen  microbes 
inside  the  white  corpuscles  had  never  deduced  from  this  a  pro- 
tective function  :  quite  the  contrary,  for  authorities  of  the  rank 
of  Waldeyer  and  Robert  Koch  firmly  believed  that  the  microbes 
found  in  the  leucocytes  only  a  field  of  growth  and  a  means  of 
dispersion  throughout  the  body.  Haeckel,  also,  had  no  idea 
that  the  presence  of  foreign  particles  in  the  amoeboid  cells  was 
the  result  of  an  active  engulfing  process.  The  development  of 
the  phagocytic  doctrine  brought  it  into  opposition  to  the 
humoral  theory,  which  was  sustained  under  the  most  varied 
forms  by  the  most  celebrated  supporters.  As  with  many  other 
doctrines  which  have  eventually  been  admitted  as  scientific 
truths,  the  doctrine  of  phagocytosis  was  revolutionary  in  con- 
ception and  had  to  conquer  by  main  force. 

It  originated  in  zoology  and  is  a  result  of  the  comparative 
method.  From  the  study  of  the  biology  of  organisms  low  in 
the  scale  of  life,  stage  by  stage  it  gained  the  field  of  medicine. 
These  stages  we  have  indicated  in  the  observations  and  ex- 
periments already  described  in  connection  with  the  digestive 
activity  of  the  mesodermic  cells,  intracellular  digestion  in 
general,  the  reaction  of  Bipinnaria  to  the  introduction  of 
splinters,  the  diseases  of  such  lower  animals  as  are  transparent 


190  MICROBES   AND  TOXINS 

and  suitable  for  observation  in  the  living  condition,  e.g., 
Daphnia,  and  finally  the  infectious  diseases  of  animals  and  man. 
"  I  have  sought  to  develop  the  conception  that  the  intracellular 
digestion  found  in  unicellular  organisms  and  in  many 
invertebrates  has  been  transmitted  by  heredity  to  the  higher 
animals  and  in  them  has  become  fixed  and  preserved  in  the 
amoeboid  cells  of  mesodermic  origin."  Phagocytosis  is  in 
harmony  with  the  Darwinian  principles  of  evolution  among 
living  beings. 

The  essential  fact  of  immunity  is  the  intracellular  absorption 
and  digestion  of  microbes  and  probably  of  toxins  under 
precisely  the  same  conditions  as  in  the  absorption  and  digestion 
of  cellular  elements  and  albuminoid  fluids  in  general  when 
introduced  into  the  body. 

The  general  laws  are  the  same  whether  it  is  a  question  of  the 
absorption  of  extravasated  blood  after  a  wound  or  an  internal 
haemorrhage,  or  of  blood  corpuscles  injected  into  the  peritoneal 
cavity  of  a  guinea-pig  ;  whether  one  is  dealing  with  cells  so 
diverse  as  spermatozoa  or  epithelial  cells,  injected  into  the 
peritoneum  of  a  foreign  species,  or  with  complex  albuminoid 
fluids  such  as  blood-serum,  milk,  egg-albumin,  or  finally  with 
the  bodies  or  toxins  of  bacteria.  Laying  aside  the  historical 
development  let  us  now  attack  the  mass  of  facts  accumulated  on 
the  subject  of  immunity. 

Taking  a  general  view  of  the  observations  and  interpretations 
which  are  multiplying  every  day  but  are  far  from  being 
universally  clear  or  certain,  two  points  of  view  are  continually 
being  opposed  to  each  other,  the  activity  of  the  cells  and  the 
activity  of  the  body-fluids ;  the  cellular  theory  and  the  humoral 
theory  of  immunity. 

The  supporters  of  the  cell  theory  do  not  deny  the  participa- 
tion of  the  body-fluids  separated,  more  or  less  artificially,  from 
the  cells,  /.*.,  the  phagocytes ;  but  they  maintain  that  the  cells 
are  the  primary  and  principal  agents,  the  humoral  properties 
being  secretions  or  excretions  of  the  phagocytes,  and  the  final 
stage  in  the  destruction  of  the  microbes  being  digestion  in  the 
interior  of  the  phagocytes. 


IMMUNITY 


191 


The  supporters  of  the  humoral  theory  consider  that  in 
immunity  the  body-fluids  (serum,  exudates,  &c.)  possess  or 
acquire  destructive  properties  independent  of  the  cells;  that 
there  is  a  non-phagocytic  destruction  of  bacteria  (and  poisons), 
and  that  even  when  this  destruction  appears  to  be  completed 
in  the  interior  of  the  leucocytes,  the  role  of  the  latter  is  limited 
to  seizing  and  absorbing  bacteria  already  killed. 

Of  course  the  antagonism  between  these  two  standpoints  is 
not  irreconcilable,  and  intermediate  theories  exist.  The 
priority  of  the  cells,  however,  as  compared  with  the  fluids 


FIG.  64. — Intracellular  (phagocytic)  diges- 
tion in  an  intestinal  cell  of  Planaria. 
(Metchnikoff.) 


FIG.  63.  —  Phagocytosis  o 
the  red  corpuscles  of  the 
goose  by  the  phagocytes 
of  the  snail.  (Metch- 
nikoff. ) 


independent  of  the  cells,  is  still  the  subject  of  dispute;  no 
theory  succeeds  in  explaining  the  facts  of  immunity  without 
acknowledging  the  activity  of  the  phagocytes  and  the  im- 
portance of  intracellular  digestion. 

The  humoral  theory  first  took  the  field  with  claims  or 
aspirations  to  be  a  "  chemical "  theory,  when  some  at  least  of 
the  phenomena  of  immunity  were  successfully  reproduced  out- 


192 


MICROBES   AND  TOXINS 


side  the  body  in  the  test-tube.  It  must  be  carefully  kept  in 
view  that  all  the  most  plausible  arguments  in  its  favour  are 
supplied  by  "  in  vitro "  experiments.  It  is  a  laudable 
tendency  to  attempt  to  reduce  biological  phenomena  to  a 
mechanism  reproducible  at  will,  but  it  must  not  be  allowed  to 
distort  the  facts  of  nature.  The  study  of  immunity  is,  above 

all,  the  study  of  an  infected 
body  defending  itself.  We 
know  neither  the  nature 
nor  the  composition  of 
the  substances  concerned, 
albuminoid  or  otherwise : 
we  are  not  even  always  sure 
that  the  substances  posUr 
lated  really  exist.  Too 
often  we  yield  to  the  ten- 
dency to  describe  as  sub- 
stances what  we  only 
observe  as  functions ;  these 
functions  have  been  sym- 
bolised by  names  and  by 
signs,  and  some  have  even 

come  to  see  in  them  actual  things  and  things  with  an  actual 
shape.  It  cannot  be  denied  that  at  present,  while  we  are 
still  awaiting  the  advances  so  generally  longed  for,  vitalism 
(in  the  sense  in  which  certain  critics  of  the  phagocytic  theory 
employ  the  word)  represents  the  most  realistic  conception. 

Metchnikoff  has  therefore  never  ceased  to  recall  and  empha- 
size the  differences  which  separate  the  corresponding  (one 
cannot  say  "  the  same  ")  phenomena  "  in  vivo  "  and  "  in  vitro  "  : 
not  that  he  disputes  the  importance  of  the  latter,  but  to  empha- 
size the  necessity  of  associating  them  always  with  the  phenomena 
in  the  living  animal.  Experiment  ought  always  to  deal  as  much 
as  possible  with  the  living  creature  itself.  The  cells  and  the 
body-fluids  are  hardly  to  be  treated  as  substances  capable  of 
preservation  in  bottles,  and  this  fact  we  will  have  occasion  to 
recall  more  than  once. 


FIG.  65. — Phagocytes  taking  up  spores 
of  the  tetanus  bacillus  (heated). 


IMMUNITY  193 

Antigens  and  Antibodies. — The  cells  which  play  a 
part  in  immunity  are  known  :  they  are  the  phagocytes,  the 
micro-  and  macro-phages.  The  humoral  properties  correspond 
to  what  are  known  as  antibodies. 

The  antibodies  are  the  products  (substances  or  properties)  of 
a  reaction  of  the  body  towards  a  natural  or  artificial  introduc- 
tion into  it  of  certain  foreign  substances,  bacteria  and  their 
poisons,  vegetable  poisons  of  other  kinds,  and  various  albumi- 
noids all  known  by  the  name  of  antigens.  The  exact  definition 
of  an  antigen  is  its  capacity  of  exciting  in  the  injected  (or 
infected)  body  the  production  of  an  antibody. 

The  discovery  of  the  antibodies  was  so  much  more  a  splendid 
biological  acquisition  in  that  its  practical  importance  was  at 
least  equal  to  its  theoretical.  The  first  antibodies  studied  were 
the  antitoxins  of  diphtheria  and  tetanus.  The  discovery  of  the 
diphtheria  toxin  by  Roux  led  to  the  discovery  of  its  antitoxin 
in  the  hands  of  Behring.  The  neutralisation  in  an  ordinary 
test-tube  of  a  toxin  by  an  antitoxin  was  one  of  the  first  and 
most  brilliant  "t'n  vitro"  experiments  in  immunity.  It  might 
certainly  seem  that  this  neutralisation  could  take  place  equally 
simply  in  the  living  animal  with  no  intervention  of  the  cells,  but 
like  a  chemical  combination. 

When  an  animal  of  species  A  is  injected  with  the  red  blood 
corpuscles  of  an  animal  of  species  B,  the  serum  of  the  former 
acquires  the  property  of  dissolving  the  globules  of  the  other 
species :  it  becomes  haemolytic  and  the  prepared  animal  is 
said  to  have  developed  a  hamotysin.  When  the  body  is 
vaccinated  against  the  typhoid  bacillus,  the  serum  acquires 
the  property  of  agglutinating  a  homogeneous  suspension  of 
typhoid  bacilli :  it  is  said  to  have  produced  an  agglutinin. 
The  serum  of  an  animal  A  which  has  been  injected  with 
the  blood  or  the  serum  of  an  animal  B  of  a  different  species 
forms  a  precipitate  when  to  it  there  is  added  a  little  of  the 
serum  B  ;  there  is  said  to  have  been  developed  a  precipitin  to  B. 

Haemolysins,  agglutinins,  and  precipitins  are  the  antibodies 
of  which  the  blood  corpuscles,  the  bacteria,  and  the  serum- 
proteins  are  the  antigens. 

o 


194  MICROBES  AND  TOXINS 

Complement  or  Alexine. — Before  the  discovery  of  the 
antibodies,  at  the  time  when  attempts  were  being  made  to 
transfuse  human  patients  with  the  blood  of  other  mammals,  in 
particular  of  the  sheep,  it  had  been  noticed  that  the  normal 
blood-serum  of  certain  animals  destroys  the  red  corpuscles  of 
other  species  :  Buchner  attributed  this  property  to  a  defensive 
substance  which  he  called  alexine.  Its  chemical  composition 
is  unknown  ;  it  is  thought  to  be  an  albuminoid  substance  :  it  is 
known  to  disappear  from  serum  on  dialysis  and  to  act  after  a 
period  of  inactivity  or  incubation ;  it  is  destroyed  by  tempera- 
tures about  56°  C.,  and  only  acts  in  presence  of  salts.  Buchner 
classes  it  along  with  the  digestive  ferments. 

The  serum  of  many  animal  species,  expressed  from  the  blood 
after  coagulation,  possesses  the  property  of  killing  in  vitro  many 
infective  bacteria  without  any  apparent  assistance  from  the 
body-cells  :  this  destructive  action  is  similarly  attributed  to  the 
alexine  or  complement,  and  it  is  this  bactericidal  property 
which  represents  the  simplest  and  crudest  fact  on  which  the 
whole  structure  of  the  humoral  theory  of  immunity  has  been 
reared. 

The  Two  Substances. — Bordet  showed  in  1895  that  the 
serum  of  an  immunized  animal  contains  two  substances,  or 
rather  two  functions.  Take  a  guinea-pig  which  has  received 
intraperitoneally  spaced  injections  of  cholera  vibrios :  its  serum 
now  destroys  these  in  vitro^  and  ,is  said  to  be  bacteriolytic. 
Heated  to  56°  C.,  it  loses  this  property,  but  on  the  addition  of 
a  little  fresh  serum,  recovers  it.  The  bacteriolytic  action  thus 
demands  primarily  a  substance  present  in  the  fresh  normal 
serum  of  any  animal  species  (non-immunized),  a  substance 
which  heating  to  56°  C.  destroys  :  it  is  the  alexine  or  comple- 
ment. But  it  demands  also  another  substance  present  in  the 
serum  of  treated  animals  and  absent  in  normal  serum,  which  is 
not  destroyed  by  heating  under  65°  C. 

Bacteriolysis  (the  destruction  of  bacteria  by  serum)  is  thus 
due  to  the  collaboration  of  two  functions,  by  custom  regarded 
as  two  substances.  One,  thermolabile,  is  the  alexine  or 
complement  and  exists  in  all  freshly  prepared  normal  sera. 


IMMUNITY  195 

The  other,  thermostable,  only  exists  (with  rare  exceptions)  in 
treated  immunized  animals :  in  the  example  given  above  it 
makes  the  cholera  vibrio  susceptible  to  the  action  of  the 
alexine  or  complement,  or  it  may  be  said  to  fix  this  to  the 
bacteria,  or,  finally,  it  may  be  regarded  as  forming  the  connect- 
ing link  between  the  alexine  or  complement  and  the  vibrios. 
It  has  been  called  by  Bordet  the  '  sensibilisatrice? 1 

The  idea  of  two  substances  is  not  a  theory,  as  Bordet 
remarks  with  great  justice;  there  is  nothing  hypothetical  in 
it.  It  is  simply  putting  in  words  the  facts  observed,  in 
particular  those  of  re-activation. 

Haemolysis,  agglutination,  precipitation,  in  a  word,  all  the 
reactions  in  which  antibodies  play  a  part,  proceed  in  the  same 
fashion  and  Bordet's  discoveries  have  thus  a  general  applica- 
tion. 

A  fundamental  experiment  performed  by  Ehrlich  and 
Morgenroth  completes  Bordet's  observations.  The  immune 
body  becomes  fixed  on  the  antigens  (bacteria  or  blood 
corpuscles)  without  producing  in  them  any  visible  modification, 
so  that  eventually  the  fluid  containing  it  is  completely  robbed 
of  immune  body;  this  fixation  prepares  the  way  for  the 
destruction  of  the  corpuscles  or  the  bacteria,  but  this  latter  does 
not  occur  till  the  complement  is  added.  On  the  other  hand, 
the  complement  does  not  fix  itself  to  the  antigen  when  alone, 
but  only  by  the  intermediation  of  the  immune  body. 

This  collaboration  of  two  substances  or  ferments  in  one 
complex  physiological  process  is  not  the  sole  example  in  biology. 
The  digestive  ferments  of  the  pancreas,  amylase,  saponase, 
and  especially  trypsin,  fail  to  exert  their  full  activity  without  the 
collaboration  of  the  enterokinase,  a  ferment  secreted  in  the 
juice  of  the  small  intestine  not  by  the  cells  and  glands  of  the 
mucous  membrane  itself,  but  by  the  lymphoid  tissue,  which  is 
composed  of  white  corpuscles  :  the  importance  of  this  fact  will 
be  seen  (Pawlow;  Chepowalnikoff ;  Delezenne). 

1  The  alexine  is  called  by  Ehrlich  the  complement  or  complementary 
substance  ;  the  sensibilisatrice  is  often  called  the  immunizing  substance ;  the 
intermediate  substance,  immune-body  or  amboceptor. 

O   2 


196 


MICROBES   AND   TOXINS 


The  antibodies  quoted  may  themselves  give  rise  to  antibodies  : 
anticomplements  and  anti-immune  bodies  may  be  prepared. 

We  know  now  all  the  factors  which  come  into  play  in  the 
phenomena  of  immunity :  on  the  one  hand  cells,  the  phagocytes  ; 
on  the  other  the  body  fluids,  containing  ferments  the  actions  of 
which  supplement  each  other,  the  immune  body  and  the  com- 
plements ;  we  know  also  that  the  experiments  of  bacteriolysis 
and  haemolysis  in  vitro  appear  to  indicate  that  the  chief 
phenomena  of  immunity  are  independent  of  the  phagocytic 
cells.  Let  us  now  examine  the  problem  more  closely  and  see 
if  phagocytosis  stands  the  test  brought  against  it  by  the  humoral 
theories. 

PHAGOCYTIC  IMMUNITY 

In  every  case  in  which  the  body  possesses  immunity  the 
bacteria  against  which  immunity  exists  are  devoured  by  the 
phagocytes,  which  collect  in  crowds,  incorporate,  and  digest 
them.  "  Looked  at  from  this  standpoint  immunity  becomes  a 
phenomenon  much  more  general  than  a  mere  resistance  of  the 
body  to  infectious  disease."  On  ultimate  examination  it 

reduces  itself  to 
a  phenomenon  of 
cellular  suscepti- 
bility, of  chemio- 
tactic  influences, 
and  of  intracellular 
digestion,  Immu- 
nity is  a  phenome- 
non of  digestion. 

Phagoc  ytosis 
can  be  directly 
observed  in  many 

(Metchnikoff.)  cases    of    natural 

immunity:    the 

disease  of  Daphnia  presents  one  of  the  simplest  and  most 
typical  examples,  and  similar  ones  have  been  observed  among 
other  invertebrates.  Among  the  vertebrates  the  frog  owes 


FIG.  66.  —  Phagocy-    FIG.    67.  —  Microphage  of 
tosis  of  anthrax  the  rat  full  of  anthrax 

bacilli     by     the  bacilli.    (Metchnikoff.) 

macrophages    of 
the    rat's    liver. 


IMMUNITY  197 

its  resistance  towards  the  anthrax  bacillus  to  phagocytosis,  the 
same  bacillus  growing  excellently  in  the  body-fluids  deprived 
of  cells. 

Similarly,  the  anthrax  bacillus  grows  very  well  in  the  body  of 
a  fowl,  although  the  fowl  is  very  resistant  to  inoculation.  The 
effect  of  cold  in  rendering  it  susceptible  (the  famous  experiment 
of  Pasteur)  is  to  be  ascribed  to  a  benumbing  of  the  phagocytes. 
In  the  case  of  the  dog  resistant  to  anthrax,  or  of  the  guinea-pig  to 
the  spirillum  of  relapsing  fever,  or  to  the  cholera  vibrio  injected 
in  small  dose  in  the  peritoneum,  the  engulfment  and  digestion 
of  the  microbes  by  the  phagocytes  are  visible  facts  and  the 
figures  appended  are  better  than  any  description. 

On  the  other  hand,  the  bacteria  cannot  be  said  to  be  expelled 
from  the  body  through  the  various  excretory  organs.  They 
are  never  found  in  the  urine,  provided  the  kidney  filtering 
action  is  intact.  They  are  never  found  in  the  sweat  unless  by 
faulty  technique  a  little  infected  blood  gets  mixed  with  it. 

There  is  no  digestion,  even  intracellular,  without  digestive 
ferments.  Under  the  microscope  the  digestion  of  the  ingested 
microbes  can  be  seen  going  on  in  the  digestive  vacuoles  of  the 
phagocytes,  and  by  means  of  the  dye,  neutral  red,  the  acidity 
of  the  part  in  which  digestion  is  proceeding  is  equally  easy 
to  demonstrate,  as  in  the  case  of  the  digestive  vacuoles  of  a 
myxomycete  or  an  amoeba,  or  as  in  the  intestinal  cells  of 
Planarians  or  Actinians.  Metchnikoff  considers  that  there  are 
two  varieties  of  leucocytic  digestive  ferments  corresponding  to 
the  two  great  groups  of  phagocytes,  the  macrophages,  which 
digest  chiefly  the  cellular  elements  and  the  bacteria  of  chronic 
infections  such  as  the  tubercle  bacillus,  and  the  microphages 
which  digest  chiefly  bacteria.  They  can  be  obtained  by  making 
extracts  of  those  organs  which  are  rich  in  phagocytes,  the 
lymphatic  glands,  the  spleen,  and  the  bone-marrow.  In  natural 
immunity  the  digestive  ferment  of  the  leucocytes  is  simply  the 
complement. 

There  have  been,  however,  many  disputes  regarding  this 
point  and  regarding  the  origin  of  complement.  Certain 
observers  have  recently  maintained  that  the  complement  has 


198  MICROBES   AND   TOXINS 

nothing  to  do  with  the  white  corpuscles.  They  have  made 
extracts  of  leucocytic  exudates  withdrawn  from  the  body,  and 
shown  that  these  extracts  were  either  without  bactericidal 
power  or  that  the  bactericidal  substance  they  contained 
possessed  properties  quite  different  from  those  of  complement. 

It  is  true  that  in  extracts  of  white  corpuscles  prepared 
by  maceration  or  freezing  no  complement  can  be  detected 
capable  of  destroying  bacteria,  but  such  experiments  do  not 
prove  that  the  production  of  complement  by  these  is 
impossible.  The  complement  may  easily  be  lost  in  the  course 
of  the  maceration  and  freezing,  rather  brutal  processes  in  any 
case  for  living  cells.  It  is  also  possible  that  the  complement 
may  be  neutralised  by  some  antagonistic  substance  contained 
in  the  leucocyte,  some  sort  of  anti-complement :  we  are 
certainly  far  from  knowing  all  the  substances  contained 
by  leucocytes.  It  is  conceivable  that  they  may  respond  to 
a  slight  injury,  received  in  the  course  of  the  manipulation 
of  the  blood,  by  discharging  into  the  surrounding  fluid  com- 
plement alone,  whereas  when  more  seriously  injured  they  may 
discharge  the  neutralizing  substance.  Later  it  will  be  seen 
that  Pfeiffer's  phenomenon  when  correctly  interpreted  supports 
this  view:  in  the  body  thoroughly  immunized  against  the 
cholera  vibrio  but  with  the  white  corpuscles  uninjured,  the 
vibrios  are  not  destroyed  by  the  body-fluids  and  are  altered 
only  in  the  interior  of  the  cells. 

According  to  Metchnikoff  the  complement  is  secreted  by  the 
phagocytes,  never  excreted,  i.e.,  poured  out  into  the  serum  or 
the  body-fluids,  so  long  as  the  phagocytic  process  remains 
normal ;  it  is  only  discharged  when  the  phagocyte  has  been 
injured  or  phagolysed,  as  this  semi-destruction  has  been  called. 
It  resembles  the  zymase  of  the  yeast  cells  of  beer,  which 
are  only  liberated  by  processes  which  break  up  the  cell.  The 
fact  is  one  of  great  importance,  as  will  be  found  again  in 
the  discussion  of  Pfeiffer's  phenomenon :  the  complement 
action  never  takes  place  outside  the  bodies  of  the  phagocytes 
except  when  there  has  been  phagolysis. 

In    acquired    immunity,    i.e.,    in    an    animal    which    has 


IMMUNITY  199 

recovered  from  an  infection  or  which  has  been  treated  in 
the  laboratory,  other  ferments  develop  which  did  not  exist, 
or  hardly  existed,  before  the  appearance  of  immunity :  these 
are  the  sensibilisatrices,  or  immune-bodies  or  amboceptors 
of  Ehrlich.  They  are  secreted  by  the  macrophages  to  some 
extent,  but  chiefly  by  the  microphages,  and  are  found  in  the 
spleen,  in  the  lymphatic  gland,  and  the  bone  marrrow  at 
a  stage  in  the  immunizing  process  when  they  are  still 
absent  from  the  blood.  They  resist  a  higher  temperature  than 
the  complement,  and  have  properties  resembling  the  ferment 
enterokinase  of  the  small  intestine.  Just  as  the  enterokinase 
prepares  fibrin  for  the  action  of  trypsin,  so  the  immune-bodies 
prepare  the  bacteria  or  other  cell-elements  for  the  action  of  the 
complement :  this  is  an  analogy  between  extracellular  and 
intracellular  digestion  which  ought  to  be  emphasized. 

There  is  not  in  any  given  animal  a  series  of  different  com- 
plements :  the  complement  from  the  same  animal  performs 
indifferently  haemolysis  and  bacteriolysis,  and  dissolves  equally 
well  the  typhoid  bacillus  and  the  cholera  vibrio.  The  immune- 
bodies,  on  the  contrary,  are  specific,  being  developed  during  the 
immunization  against  the  invading  cells  (by  inoculation  or 
natural  infection). 

Complement  is  discharged  into  the  fluids  bathing  the  phago- 
cytes only  when  phagolysis  has  occurred  :  the  immune-bodies, 
on  the  other  hand,  are  readily  excreted  by  the  phagocytes  ;  they 
resemble,  not  zymase,  which  is  firmly  bound  to  the  protoplasm 
of  the  yeast  cell,  but  sucrase,  which  is  easily  discharged. 
Recovery  or  inoculation  does  not  increase  the  quantity  of  com- 
plement, but  greatly  develops  the  quantity  of  immune-body. 
In  natural  immunity  the  presence  of  immune- body  is  difficult 
to  demonstrate,  probably  because  there  is  little  of  it  in  existence, 
and  what  there  is  is  contained  in  the  phagocytes ;  but  in 
acquired  immunity  immune-body  is  abundant  and  is  found,  not 
only  in  the  plasma  and  serum  of  the  blood,  but  in  exudates  and 
cedematous  fluids.  Examples  of  acquired  immunity  exist  in 
which  the  body  remains  poor  in  immune-body  and  in  which 
the  body-fluids  entirely  lack  it ;  in  these  it  is  necessary  to 


200  MICROBES   AND   TOXINS 

assume  that  the  action  goes  on  in  the  interior  of  the 
phagocytes. 

To  refute  the  old  opinion  that  the  leucocytes  form  a  good  cul- 
ture medium  for  bacteria  and  serve  as  convenient  vehicles  for 
them,  it  has  been  necessary  to  show  that  in  the  phagocytes  the 
bacteria  die  and  are  digested.  To  refute  the  opinion  that  the 
phagocytes  simply  incorporate  bacteria  already  damaged  or 
killed  by  other  (humoral)  actions,  it  has  been  necessary  to 
show  that  bacteria  are  ingested  in  a  living  and  virulent  condition : 
as  a  matter  of  fact  living  motile  bacilli  (B.  pyocyaneus}  can  be 
seen  in  the  interior  of  a  frog's  leucocyte.  Fatal  anthrax  can  be 
produced  in  a  guinea-pig  by  inoculating  anthrax  bacilli  already 
engulfed  by  the  phagocytes  of  a  frog  :  it  is  only  necessary  not 
to  wait  too  long,  or  digestion  in  the  leucocyte  may  be  completed. 
Pasteur  noted  that  it  was  perfectly  easy  to  kill  the  fowl  and  the 
rabbit  by  inoculating  them  with  bacilli  of  fowl  cholera  already 
incorporated  by  the  leucocytes  of  the  refractory  guinea-pig. 
The  bacteria  attacked  by  the  phagocytes  are  therefore 
thoroughly  alive  and  virulent. 

If  we  take  an  animal  immunised  against  vibrios  and  inocu- 
late it  with  the  same  microbes  against  which  it  is  immune ;  if 
we  withdraw  now  a  drop  of  the  exudate  provoked  by  the 
inoculation  and  make  of  it  a  hanging  drop  in  a  sealed  chamber 
at  incubator  temperature,  we  find  that  the  phagocytes  thus 
withdrawn  from  the  body  promptly  die  and  the  bacteria  grow 
in  their  interior  as  if  in  culture.  But  from  the  phagocytes 
withdrawn  from  the  animal  a  little  later  no  such  culture  can  be 
obtained ;  the  phagocytes  have  had  time  to  digest  the  bacteria. 
No  better  conception  can  be  furnished  than  this  of  the  life  and 
death  of  bacteria  in  the  phagocytes. 

It  has  been  said  that  the  body-fluids  have  already  attenuated 
the  virulence  of  the  microbe  before  its  capture  by  the  phagocyte. 
If  such  previous  attenuation  exists  their  remains  still  to  be  settled 
whether  it  is  due  to  a  cellular  or  to  a  humoral  action  :  in  any 
case  it  is  far  from  being  the  rule.  In  Charrin  and  Roger's 
experiments  the  streptococcus,  the  pneumococcus,  and  the 
pyocyaneus  bacillus  grown  in  the  serum  of  an  immunized 


IMMUNITY  201 

animal  no  longer  killed  fresh  animals  :  but  this  was  due  to  the 
fact  that  they  were  saturated  with  the  immune  serum  which 
contained  immune-bodies  :  deprived  of  these  by  thorough 
washing,  they  regained  their  original  virulence. 

It  has  further  been  said  that  the  microbes  act  through  their 
toxins  and  that  the  body-fluids  of  an  immunized  animal  begin 
by  neutralizing  this  toxicity,  after  which  the  bacteria  fall  easy 
victims  to  the  phagocytes.  But  if  this  were  the  case  why  should 
there  be  such  profound  differences  between  the  immunity 
towards  the  microbes  and  that  towards  their  toxins?  Why 
should  there  be  in  animals  immunized  against  the  bacillus 
pyocyaneus  or  the  cholera  vibrio  a  complete  resistance  to  infec- 
tion with  these  microbes  along  with  a  susceptibility  to  the 
toxins  equal  to  that  of  a  fresh  animal  ? 

In  all  these  objections  to  phagocytic  immunity  it  is  always 
the  idea  of  a  direct  primitive  action  of  the  body- fluids  which 
appears,  the  idea  of  the  humoral  theory.  Since  it  is  admitted 
that  the  immune-bodies  circulate  in  the  plasma  whereas 
according  to  Metchnikoff  the  complement  remains  in  the 
phagocytes,  since  the  phagocytic  theory  maintains  that  no 
excretion  of  complement  occurs  without  phagolysis,  it  ought 
certainly  to  have  been  on  this  point  that  the  humoral  theory 
should  have  made  its  attacks.  This  is  the  central  point  on 
which  turns  the  whole  question  :  if  without  phagolysis  there  is  no 
extracellular  destruction  of  microbes  in  an  immunised  animal, 
when  destruction  takes  place  outside  the  phagocytes  it  means 
there  has  been  an  abnormal  lesion,  a  phenomenon  unlikely  to 
occur  spontaneously  in  nature  and  probably  only  an  experi- 
mental accident ;  it  was  the  celebrated  experiment  of  PfeifTer 
which  threw  the  question  into  prominence. 

PFEIFFER'S  PHENOMENON  AND  THE  HUMORAL  THEORY 

It  was  the  following  experiment  of  Behring  and  Nissen  which, 
after  the  primary  investigations  of  Fliigge,  Nuttall  and  Buchner, 
seemed  best  to  explain  immunity  by  the  bactericidal  power  of 
the  body-fluids ;  the  serum  of  guinea-pigs  well  vaccinated  against 


MICROBES  AND  TOXINS 

the  vibrio  Metchnikowii,  a  cholera  vibrio,  becomes  much  more 
powerfully  bactericidal  than  the  serum  of  fresh  guinea-pigs.1 

It  is  easy  to  immunize  guinea-pigs  against  lethal  doses  of 
the  cholera  vibrio  injected  intraperitoneally.  Pfeiffer  took  a 
guinea-pig  thus  prepared,  injected  into  it  a  certain  quantity  of 
vibrios,  and  then  abstracted  from  its  peritoneum  a  little  of  the 
exudate.  He  found  that  after  a  few  minutes  the  vibrios  had 
almost  entirely  disappeared  from  the  peritoneum  ;  they  had 
been  transformed  into  granules,  the  first  stage  of  destruction, 
the  "commas"  turning  into  "dots."  Later  these  granules 
dissolved  in  the  peritoneal  fluid  like  a  piece  of  sugar  in 
water.  The  same  phenomena  were  observed  when  the  vibrios 
were  injected  along  with  immune  guinea-pig  serum  into  the 
peritoneum  of  a  fresh  guinea-pig. 

Pfeiffer' s  interpretation  was  that  in  the  immunized  body  the 
bacteria  are  destroyed  directly  by  the  body-fluids  without  the 
intervention  of  the  leucocytes. 

Such  then  is  Pfeiffer's  phenomenon,  so  long  discussed  and 
for  long  a  sort  of  touchstone  in  the  two  immunity  doctrines. 
Metchnikoff  and  his  pupils  have  subjected  it  to  merciless 
criticism.  First  of  all  they  showed  that  the  granule  formation 
takes  place  also  outside  the  body  when  the  vibrios  are  mixed 
with  a  little  fresh  serum  from  an  immunised  guinea-pig,  or  even 
when  to  the  same  serum,  which  from  age  or  heating  has  lost 
its  complement,  a  little  fresh  peritoneal  fluid  is  added.  (It 
was,  in  fact,  while  repeating  Pfeiffer's  experiment  that  Bordet 
discovered  the  two  substances  in  the  serum  of  immune  guinea- 
pigs).  In  the  test-tube,  as  in  the  peritoneum,  the  vibrios  fall 
victim  to  the  action  of  the  complement  through  the  inter- 
mediation of  the  immune-body. 

Since  the  granule  formation  is  due  to  the  combined  action 
of  the  two  substances,  and  since  we  know  that  the  leucocytes 
do  not  readily  shed  their  complement,  which  is  therefore 

1  It  must  be  said  in  this  connection  that  the  experiment  was  insufficient 
to  permit  a  general  conclusion  on  the  nature  of  acquired  immunity  ;  a 
similar  experiment  with  other  bacteria  gives  a  different  result  and  even  the 
vibrio  itself  when  injected  into  an  immunised  animal  remains  alive  in  its 
body  for  several  days. 


IMMUNITY 


rarely  found  in  the  normal  body-fluids,  this  destructive  action 
ought  not  to  take  place  in  these  nor  in  any  position  except  the 
peritoneum.  As  a  matter  of  fact,  if  the  immune  guinea-pig  is 
injected  under  the  skin,  in  the  anterior  chamber  of  the  eye,  or 
in  the  fluid  of  a  passive  oedema,  the  phenomenon  does  not 
take  place :  the  immune-body  is  present,  but  not  the  comple- 


FIG.  68. — Cholera  vibrios  phagocyted 
by  a  macrophage  of  the  guinea-pig 
and  not  yet  transformed  into 
granules.  ( Metchnikoff.) 


FIG.  69. — Cholera  vibrios 
phagocyted  by  a 
microphage  of  the 
guinea-pig  and 
turned  into  granules. 
(Metchnikoff. ) 


ment.  When  this  latter  is  added  in  the  form  of  a  little  fresh 
serum,  the  transformation  into  granules  occurs. 

Why  then  does  the  phenomenon  occur  in  the  peritoneum  if 
the  complement  remains  inside  the  leucocytes  ?  It  is  because 
the  mere  act  of  intraperitoneal  injection  produces  phagolysis. 
The  injection  of  any  liquid  into  the  peritoneum,  water  or 
nutrient  broth,  for  example,  destroys  some  at  least  of  the 
leucocytes  which  are  found  in  it :  they  discharge,  as  is  known, 
one  ferment  under  these  conditions,  that  which  produces 
coagulation  of  the  blood  ;  in  the  same  way,  they  discharge  this 
other  ferment,  the  complement,  which  acts  upon  the  sensitized 
vibrio. 

If  then  this  initial  phagolysis  could  be  prevented,  the 
phenomenon  of  Pfeiffer  would  also  fail.  Experiment  has 
proved  this :  by  injecting  into  the  peritoneum  sterile  broth, 
freshly  prepared  and  tepid,  the  leucocytes  are  rendered  much 
less  sensitive  to  a  succeeding  injection,  and  in  the  peritoneum 
thus  habituated,  thus  "  prepared,"  Pfeiffer's  phenomenon  does 


204  MICROBES   AND   TOXINS 

not  occur.  When  phagolysis  is  thus  prevented,  the  vibrios  in  all 
the  variations  of  the  experiment  possible  fail  to  be  transformed 
into  granules  but  disappear  by  digestion  in  the  interior  of  the 
phagocytes.  If  we  take  a  guinea-pig  strongly  immunized 
against  the  vibrio  of  cholera  and  inject  these  bacteria  directly 
into  its  circulation  by  the  jugular  vein,  we  find  half-an- 
hour  after  no  granule  transformation  in  the  circulating  blood 
the  vibrios  retain  their  shape  and  are  to  be  seen  inside  the 
leucocytes.  No  phagolysis  has  occurred,  accordingly  no 
Pfeiffer's  phenomenon,  and  no  extra-cellular  destruction  by  the 
body-fluids. 

The  resistance  of  the  immunized  guinea-pig  depends  so 
much  upon  phagocytosis,  that  if  the  leucocytic  activity  is  paralysed 
by  means  of  a  dose  of  opium,  the  animal  succumbs  to  a 
smaller  dose  of  vibrios  than  is  necessary  to  kill  the  non- 
narcotized  animal. 

The  destruction  after  phagolysis,  Pfeiffer's  phenomenon,  is 
not  even  a  general  fact.  It  is  true  of  the  cholera  vibrio,  a 
fragile  bacterium,  but  even  with  the  typhoid  bacillus,  also 
comparatively  fragile,  it  is  only  a  modified  Pfeiffer's  phen- 
omenon which  occurs.  With  the  bacillus  pyocyaneus  there  is  still 
greater  resistance,  and  greater  still  with  the  bacilli  of  swine- 
erysipelas  and  anthrax.  In  those  cases  in  which  the  humoral 
action  is  imperceptible ',  phagocytosis  is  active  and  constant. 

Opsonins  and  Bacteriotropins. — There  exist  more 
recent  theories  which,  while  recognizing  the  action  of  the 
phagocytes,  attribute  to  the  body-fluids  an  important  part  in 
immunity :  they  are  said  to  prepare  the  bacteria  for  phagocytic 
digestion.  These  preparatory  substances  or  actions  are  the 
opsonins  of  Wright  and  the  bacteriotropins  of  Neufeld.1 

1  In  their  experiments  these  workers  have  pursued  the  same  general 
method  ;  they  have  studied  the  phagocytosis  occurring  with  leucocytes 
withdrawn  from  the  body  and  suspended  in  glass  tubes,  i.e.,  the  phago- 
cytosis in  vitro  already  studied  in  the  old  experiments  of  Denys  and 
Leclef.  Metchnikoff  himself  did  not  fail  to  compare  phagocytosis  in 
vivo  and  in  vitro,  having  observed  the  incorporation  of  the  anthrax 
bacillus  by  leucocytes  suspended  in  urine  and  in  aqueous  humour  which  had 
been  boiled  and  thus  deprived  of  all  antibodies. 


IMMUNITY  205 

According  to  Wright  the  opsonins  are  the  principal  and 
determining  cause  of  phagocytosis,  the  act  of  incorporation  by 
the  leucocytes  being  only  the  final  concluding  operation. 
Opsonins,  being  the  essential  factor  in  immunity,  ought  not  to 
be  present  in  the  serum  of  fresh  animals,  i.e.,  there  ought  not 
to  be  any  phagocytosis  without  opsonins,  no  spontaneous 
phagocytosis.  But  spontaneous  phagocytosis  is  incontestable, 
as  has  been  established  by  the  experiments  of  Metchnikoff  and 
Bordet :  it  is  only  necessary  in  in  vitro  experiments  to  allow 
enough  time  for  it  to  take  place :  and  from  the  moment  that 
the  existence  of  spontaneous  phagocytosis  is  granted  opsonins 
can  no  longer  play  the  primary  part  in  the  phagocytic 
process. 

It  is  certain  that  the  presence  of  normal  serum  favours 
phagocytosis  in  vitro  (Wright  and  Douglas,  etc.) :  the  serum 
acts  on  the  bacteria,  which  are  capable  of  fixing  certain  of  its 
elements.  Are  the  opsonins  substances  or  properties  new  and 
unknown  before  Wright's  researches  ?  Numerous  experiments 
ascribe  to  the  opsonins  of  normal  serum  the  same  properties  as 
characterize  the  complement.  They  are  products  of  the 
leucocytes. 

In  the  serum  of  immunized  animals,  which  favours  phagocytosis 
much  more  actively  than  normal  serum,  the  opsonins  are  not 
to  be  distinguished  from  the  immune-body :  they  can  be  used 
for  the  same  re-activation  experiments  (Levaditi)  and  they  also 
are  products  of  the  phagocytes. 

The  bacteriotropins  of  Neufeld  are  considered  by  the 
majority  of  workers  as  being  equivalent  to  the  opsonins  of 
immune  serum  and  to  the  immune-body. 

There  is  no  reason  to  deny  these  actions  which  favour 
phagocytosis.  The  work  on  opsonins  and  bacteriotropins  is 
simply,  to  use  Ehrlich's  expression,  a  new  flowering  of  the 
phagocytic  doctrine.  In  Wright's  and  Neufeld's  experiments  it 
is  chiefly  the  experimental  method  which  Metchnikoff  criticises. 
Leucocytes  taken  from  the  body,  washed  and  in  a  different 
surrounding  medium,  can  no  longer  accurately  represent  the 
phenomena  occurring  in  the  living  body.  The  conditions  are 


206  MICROBES   AND   TOXINS 

abnormal  and  yet  one  knows  that  it  is  only  in  abnormal  conditions 
that  the  leucocytes  discharge  complement.  "  The  least  change 
in  the  salt  content  of  the  surrounding  fluid  is  sufficient  to 
modify  notably  the  phagocytosis.  The  leucocytes,  of  patients 
suffering  from  various  diseases  present  a  marked  diminution 
in  their  vital  activities.  The  destruction  of  bacteria  is 
the  work  of  phagocytes  which  are  living  and  vigorous" 
(Metchnikoff,  Nobel  Lecture).  Washing,  chilling,  maceration 
are  quite  sufficient  to  destroy  the  complement  of  the  leucocytes ; 
how  is  it  possible  to  conclude  after  such  procedures  that  the 
leucocytes  do  not  contain  the  complement  ? 

The  opponents  of  phagocytosis  declare  that  it  is  the  humoral 
properties  which  undergo  the  most  marked  increase  during 
immunization.  There  is  no  doubt  of  such  a  development  of 
bacteriotropins,  opsonins  and  immune-bodies — which  in  any 
case  are  phagocytic  products.  But  it  can  be  shown  experi- 
mentally that  the  phagocytes  are  modified  in  immunity  and 
modified  sooner  than  the  body-fluids.  Leucocytes  taken  from 
an  animal  vaccinated  against  some  microbe  and  injected  into 
a  fresh  animal  protect  the  latter  from  several  lethal  doses 
of  the  microbe,  whereas  the  leucocytes  of  a  normal  animal  fail. 
(Pettersson's  experiment).  The  white  corpuscles  of  the 
immunized  animal  supply  protective  substances  at  a  time  when 
the  blood-fluids  are  not  yet  affected :  and  it  is  owing  to  the 
leucocytes  that  the  body  remains  refractory  after  the  body 
fluids  have  already  lost  their  protective  properties  (Salimbeni). 

Serum  is  a  fluid  into  which  have  been  poured  the  ferments 
of  the  lencocytes,  the  fibrin-ferment  and  the  complement. 
Injury  to  the  leucocytes  is  necessary  before  blood  will  coagulate. 
By  very  delicate  operations  and  with  great  trouble  it  has  been 
possible  to  separate  the  blood  corpuscles  and  obtain  a 
plasma  which  remains — for  a  certain  time — incoagulable. 
Now  the  properties  of  such  plasma  are  very  different  from 
those  of  serum  :  the  leucocytic  excretions  are  absent.  It  is,  how- 
ever, so  difficult  to  obtain  a  true  plasma  identical  with  that  of 
the  circulating  blood  that  such  experiments  have  to  be  very 
carefully  analysed.  When  quickly  prepared  immediately  after 


IMMUNITY  207 

bleeding  the  plasma  contains  no  complement,  but  every 
minute  afterwards  injured  leucocytes  pour  into  it  little  by 
little  the  active  substance. 

Antibodies  and  Immunity. — There  are  innumerable 
facts  preventing  us  from  regarding  as  a  law  any  correspondence 
between  the  quantity  of  antibodies  in  the  serum  and  the 
degree  of  immunity  of  the  animal :  this  is  a  definite  proof  that 
there  is  something  else  besides  the  humoral  properties  and  that 
the  preponderating  part  is  played  by  the  cell  elements. 

The  serum  of  guinea-pigs  inoculated  against  anthrax  was 
found  by  Behring  and  Wernicke  to  be  incapable  of  protecting 
fresh  guinea-pigs  from  a  fatal  infection.  Pfeiffer  immunised 
guinea-pigs  against  the  bacterium  which  he  regards  as  the  cause 
of  influenza  in  man,  but  these  immunized  animals  did  not 
produce  a  protective  serum. 

In  protozoal  diseases  such  as  malaria,  there  seems  to  be 
immunity  in  certain  cases,  but  no  one  has  ever  demonstrated 
a  protective  property  in  the  serum.  To  take  an  example 
among  the  invertebrates,  the  larvae  of  the  rhinoceros  beetle 
(Oryctes  nasicornis]  are  immune  to  anthrax  and  in  them  the 
phagocytic  incorporation  of  injected  anthrax  bacilli  can  be 
very  well  seen  ;  yet  the  blood  fluid  of  these  larvae  forms  a 
culture  medium  equally  favourable  to  the  anthrax  bacillus,  to 
which  they  are  immune,  as  to  the  cholera  vibrio  which  produces 
in  them  a  fatal  infection. 

Again,  the  dog  is  extremely  resistant  to  anthrax,  yet  the 
anthrax  bacillus  grows  very  well  in  dog- serum ;  all  these  are 
examples  of  immunity  in  which  the  bactericidal  power  does 
not  play  a  part.  It  has  been  known  since  the  experiments  of 
Behring  that  rat's  serum  possesses  a  remarkable  destructive 
power  towards  the  anthrax  bacillus ;  now  the  rat  is  not 
immune  to  anthrax,  and  the  degree  of  immunity  which  it  does 
possess  is  due  to  the  phagocytes.  The  bactericidal  substance 
exists  in  the  leucocytes,  but  not  in  the  circulating  plasma,  nor 
in  plasma  carefully  prepared  by  Gengou's  method.  It  exists 
in  the  serum,  but  only  because  it  has  been  discharged  into 
this  by  the  leucocytes.  The  rat  is  extremely  susceptible  to 


208  MICROBES   AND  TOXINS 

anthrax  if  it  is  inoculated  with  a  very  fine  needle  so  as  not  to 
provoke  a  haemorrhage  at  the  point  of  inoculation.1 

Immunity  towards  toxins  could  be  made  to  furnish  analogous 
examples  in  abundance ;  it  will  be  discussed  in  the  next 
chapter. 

In  many  cases  Pfeiffer  has  seen  his  guinea-pigs,  which  had 
been  thoroughly  immunized  against  the  cholera  vibrio,  succumb 
to  the  injection  of  a  moderate  dose  of  vibrios.  Yet  the  serum 
of  these  guinea-pigs  was  capable  of  producing  Pfeiffer's  phe- 
nomenon.2 

Tuberculin  carefully  employed  exerts  a  favourable  effect  in 
many  tuberculous  patients,  and  leads  to  the  production  of 
antibodies  in  their  serum.  Jochmann  has  quite  recently  made 
several  observations  of  this  kind  under  the  direction  of  R.  Koch, 
and  has  sought  for  a  correspondence  between  the  appearance 
and  quantity  of  the  antibodies  and  the  resistance  of  the 
patient.  He  found  it  was  impossible  to  maintain  that  the 
presence  of  antibodies  meant  recovery.  Certain  patients  mani- 
fested great  clinical  improvement  simultaneously  with  the 
appearance  of  antibodies;  in  others,  the  improvement  was 
quite  as  great  without  the  antibodies,  while  in  other  cases  the 
appearance  of  the  antibodies  coincided  with  marked  and  fatal 
aggravation  of  the  malady.  It  is  obviously  impossible  to  draw 
any  conclusion  as  to  the  immunity  of  these  patients  from  such 
in  vitro  experiments. 

1  In  this  example  of  the  rat  there  is  no  question  of  an  antibody  produced 
by  immunization,  but  rather  of  a  want  of  agreement  between  the  natural 
immunity  of  the  animal  and  the  natural  bactericidal  power  of  the  serum . 

2  Quite   recently   Citron   has  shown  that  the  serum  of  rabbits  actively 
immunised  against  the  so-called  bacillus  of  hog-cholera  possesses  protective 
properties  for  guinea-pigs,  whereas  it  has  none  of  this  for  fresh  rabbits. 
Rabbits  prepared  with  extracts  of  the  bacilli  and  without  active  immunity 
of  their  own  (they  succumbed  to  the   test  inoculation  of  living  bacilli), 
nevertheless  furnished  a  protective  serum  for  guinea  pigs.     Choukewitch 
taking    up    this    question    again,    prepared   rabbits    by  large  intravenous 
inoculations  of  the  same  bacilli,   but  in  the  killed  condition.     One  rabbit 
of  the  lot  acquired  immunity  towards  the  living  microbes,  but  in  every  one 
of  the  series,  not  only  the  immune  individuals  but  also  the  non-immune, 
the  blood  contained  an  abundance  of  antibodies,  immune-bodies,  opsonins, 

etc It   even    contained   much    more   than   the   serum   of  rabbits 

rendered  truly  immune  by  subcutaneous  inoculations  of  virulent  bacilli. 


IMMUNITY  209 

Immunity  is  entirely  a  cellular  function  and  the  inherent 
tincture  of  "  vitalism  "  in  the  phagocytosis  doctrine  is  unavoid- 
able. 

"  The  final  phagocytic  reaction  is  represented  by  the  physical 
or  physico-chemical  processes  in  the  digestion  of  microbes, 
conducted  with  the  help  of  cytases,  and  favoured  by  the 
presence  of  immune  bodies;  in  the  resistance  to  poisons  the 
phagocytes  must  also  exert  chemical  influences.  But  before 
these  phenomena  occur,  the  phagocytes  present  activities  which 
are  purely  biological ;  such  are  the  chemiotactic  perceptions 
and  movements  directed  towards  the  threatened  spot,  the 
engulfment  of  bacteria,  and  the  absorption  of  toxins,  and 
finally  the  secretion  of  substances  utilized  in  cellular  digestion  " 
(MetchnikorT,  L Immunity  p.  390). 


CHAPTER  XI 

IMMUNITY 

Toxins  and  antitoxins— Chemical  and  physical  conceptions  of  immunity. 

Side-chain  theory — Origin  of  the  antibodies — Theory  of  chemical  equi- 
librium. 

The  physical  point  of  view  :  Bordet — Phenomena  of  absorption  or  molecular 
adhesion — Explanation  of  specificity — Analogies  between  the  reactions 
of  antibodies  and  the  reactions  of  colloids— Lipoid  actions. 

Phagocytosis  and  toxins — The  body  plays  an  essential  part — Origin  of 
antibodies  and  Wassermann's  experiments — The  phagocytes  in  their 
connection  with  mineral  poisons  and  microbial  poisons,  toxins,  and 
endotoxins. 

IT  was  the  discovery  of  antitoxins  which  inaugurated  the  study 
of  antibodies.  It  was  the  necessity  of  explaining  the  action  of 
antitoxins  on  toxins  which  gave  rise  to  the  theories  on  this 
peculiar  problem  and  on  antibodies  and  immunity  in  general. 

It  was  thought  at  first  that  in  the  immunized  animal  which 
manufactures  the  antitoxin,  as  well  as  in  the  animal  immunized 
by  the  injection  of  the  antitoxic  serum,  the  cells  play  a  part. 
Buchner  enunciated  the  hypothesis  that  the  body  produces 
antitoxin  by  transforming  the  toxin ;  he  quoted,  as  a  distant 
analogy,  the  transformation  of  one  compound  into  another  by 
polymerization.  But  it  is  difficult  to  understand  how  there 
could  be  such  a  disproportion  between  the  toxin  injected  and 
the  antitoxin  produced;  the  horse  produces,  according  to 
Knorr,  for  one  unit  of  toxin  injected  100,000  units  of  anti- 
toxin. 

Buchner  performed  a  pretty  experiment,  which  has  lost  none 
of  its  interest,  by  showing  that  after  accounting  for  the 
differences  in  weight  and  in  natural  susceptibility,  a  mixture  of 

210 


IMMUNITY  211 

tetanus  toxin  and  antitoxin  which  is  neutral  for  the  guinea-pig 
is  fatal  to  the  mouse ;  it  is  impossible  to  avoid  the  conviction 
that  the  body,  that  of  the  guinea-pig  or  of  the  mouse  in  this 
example,  counts  for  something  in  the  phenomena.  The  same 
ideas  were  maintained  by  Roux  at  the  time  of  the  discovery  of 
serotherapy. 

But,  on  the  other  hand,  the  action  of  antitoxin  on  toxin 
seemed  to  be  a  neutralization,  and  to  behave  both  in  vitro  and 
in  vivo  like  a  chemical  reaction ;  and  in  practice  it  was  found 
necessary  for  medical  purposes  to  titrate  the  sera  in  this  way 
to  measure  their  activity ;  the  ideas  of  Buchner  and  of  Roux 
were  then  laid  aside,  only  to  be  rediscovered  later;  the 
biological  phenomena  were  subordinated  as  much  as  possible 
to  quantitative  studies,  and  the  endeavour  was  made  to 
represent  the  action  of  antitoxins  on  toxins  as  a  chemical 
reaction. 

The  Side-chain  Theory. — The  best  known  chemical 
theory  for  the  action  of  antitoxins  and  antibodies  in  general 
is  that  of  Ehrlich,  which  is  currently  known  under  the  name 
of  the  "  side-chain  theory."  The  primary  idea  of  its  author 
was  to  find  in  the  facts  mutual  relations  as  much  as  possible 
fixed  and  independent  of  the  body,  and  to  eliminate  all 
"  vitalism  "  in  favour  of  exact  quantitative  work.  To  begin 
with,  he  adopted  the  method  of  in  vitro  experiment.  The 
nature  of  the  tetanus  and  diphtheria  toxins  had  been  rendered 
much  clearer  by  the  study  of  other  toxins  more  easy  to  work 
with,  such  as  haemolysins,  agglutinins,  ferments,  and  anti- 
ferments  (ricin,  abrin,  rennin,  and  antirennin,  etc.).  Pre- 
liminary experiments  in  vitro  showed  the  general  applicability 
of  the  same  laws.  Originally  Ehrlich  believed  that  the  curative 
and  preventive  action  in  vivo  was  equivalent  to  the  neutralising 
action  towards  the  diphtheria  toxin  in  vitro,  a  belief  which  later 
experiments  were  to  disturb. 

The  second  step  was  to  demonstrate  that  antitoxin  does  not 
destroy  toxin,  but  that  the  two  bodies  combine  to  form  a  com- 
pound (neutral  from  the  physiological  point  of  view),  just  as  an 
acid  and  a  base  combine  to  form  a  salt. 

P   2 


MICROBES   AND   TOXINS 

Antitoxin  does  not  destroy  toxin  because  the  two  reacting 
bodies  can  be  recovered.  For  example,  the  neurotoxin  of 
cobra  venom  resists  heating  to  68°  C.  ;  when  the  neutral  mixture 
of  venom  and  anti-venom  is  heated,  the  toxin  can  be  recovered 
if  this  is  done  during  the  ten  minutes  which  follow  the  preparations 
of  the  mixture  (Calmette's  experiment).  From  a  neutral  mixture 
of  cobra-haemolysin  and  anti-venom,  it  is  possible  to  recover  the 
haemolysin  by  the  action  of  hydrochloric  acid ;  the  recovered 
haemolysin  manifests  its  action  on  the  addition  of  the  necessary 
lecithin  (Morgenroth's  experiment).  A  mixture  of  diphtheria 
toxin  and  antitoxin  (twenty-four  hours'  contact),  harmless  to 
the  rabbit,  recovers  its  toxicity  when  treated  with  hydrochloric 
acid,  the  toxin  being  set  free  (Morgenroth  and  Willanen's 
experiment).  Heat,  nitration,  the  action  of  a  digestive  diastase, 
are  other  methods  of  dissociating  the  toxin-antitoxin  com- 
bination provided  the  intervention  takes  place  without  too 
long  delay. 

How  then  are  the  toxin  and  anti-toxin  to  be  represented  ? 
Probably  as  albuminoid  substances  of  large  molecules  capable 
of  being  represented  by  stereochemical  models  and  possessing 
a  nucleus  on  which  are  grafted  lateral  chains.  To  conceive  of 
the  action  of  the  toxin  on  a  cell,  it  is  only  necessary  to  imagine 
the  molecules  of  the  cell  protoplasm  as  containing  figures  of 
the  same  kind.  The  toxin  molecules  enter  into  relation  or 
combination  with  the  cell  molecules  by  means  of  these  atom 
groups  or  side-chains.  Ehrlich  calls  them  "  receptors " 
or  "  haptophorous  groups."  The  term  "  side-chains "  was 
borrowed  directly  from  the  chemistry  of  benzene.  Such 
stereochemical  symbols  were  introduced  into  science  by  Emil 
Fischer  to  represent  the  specific  action  of  the  ferments  :  one 
body  acts  specifically  on  another,  because  an  atom  group  of 
the  one  is  adapted  to  an  atom  group  of  the  other,  as  with  a 
key  and  the  corresponding  lock.  Thus  the  haptophorous  group 
of  a  toxin  fixes  itself  on  the  receptor  of  a  cell,  both  haptophore 
and  receptor  being  "  side-chains." 

This  in  fact  is  the  central  idea  of  the  theory,  the  same  mode 
of  chemical  action,  the  same  relations  between  receptors  and 


IMMUNITY  213 

haptophorous  groups,  conceived  after  the  manner  of  the 
reactions  of  organic  chemistry ;  and  this  molecular  stereo- 
chemistry explains  all  vital  phenomena  :  the  action  of  a  toxin 
on  the  cell,  the  action  of  antitoxin  on  toxin,  the  production  of 
antitoxins,  and  immunity  in  general. 

Finally,  since  the  same  combinations  and  linkages  go  on  in 
the  metabolic  changes  of  all  living  matter,  the  conception  of 
side-chains  becomes  a  general  theory  of  nutrition ;  so  much 
so  that,  with  all  his  chemical  language  and  mechanical  attitude 
of  thought,  Ehrlich  arrives  at  the  same  formula  as  Metchnikoff 
with  his  "  vitalistic,"  or  rather,  biological  explanation: 
immunity  is  a  function  of  nutrition. 

This  idea  has  been  Ehrlich's  guiding  principle  in  all  his 
scientific  work,  and  it  is  from  it  that  we  have  to  start  in  order 
to  arrive  at  the  principal  laws  of  immunity. 

Let  us  represent  the  protoplasm  molecule  as  possessing 
numerous  and  varied  functions,  the  agents  or  the  bases  of 
which  are  distinct  atomic  groups.  This  molecule  consists  of  a 
central  nucleus  (analogous  to  the  nucleus  of  the  aromatic 
compounds),  which  maintains  its  continuous  individuality, 
and  of  numerous  side-chains  or  receptors  which  act  towards 
the  nucleus  as  organs  of  communication  or  nutrition.  The 
primary  food-stuffs  and  the  toxins  which  circulate  in  the  blood 
and  body-fluids  have  haptophore  groups  which  are  fixed  by 
the  cell  receptors;  and  it  is  thus  that  all  the  modifications 
of  the  protoplasm  are  carried  on. 

Take  a  poison  like  the  tetanus  toxin  introduced  into  the 
body.  We  know  by  definite  experiments  that  it  is  "  fixed  " 
by  various  cells  and  in  particular  by  the  nerve-cells.  The 
toxin  molecule  is  treated  by  certain  definite  receptors  of  the 
nerve-cell  as  a  food  material  j  it  possesses  a  haptophore  group 
which  hooks  on  to  the  receptor,  but  it  possesses  also  a  toxophore 
group,  an  atom  group  which  exerts  the  toxic  action ;  it  is 
through  the  haptophore  group  that  the  toxophore  group  is  fixed 
by  the  cell  and  acts  as  a  poison. 

If  the  toxin  has  been  injected  in  sufficient  quantity, 
numerous  cell  receptors  are  occupied,  monopolized  by  the 


214  MICROBES  AND  TOXINS 

toxic  molecules;  the  cell,  deprived  of  the  use  of  these 
receptors,  has  its  functional  activity  diminished  and  its  nutrition 
threatened.  But  all  injured  protoplasm  possesses  a  reparative 
or  regenerative  power ;  the  cell  reproduces  receptors  ;  it  even 
manufactures  a  great  many  more  than  is  necessary.  To  follow 
Weigert's  dictum  the  living  matter  overstimulated  by  the  lesion 
regenerates  itself  much  beyond  its  needs ;  these  receptors 
regenerated  in  excess  are  cast  off  by  the  cell  and  pass  into  the 
body-fluids ;  and  it  is  these  which  fix  and  neutralize  a  fresh 
dose  of  toxin  injected  into  the  body. 

Antitoxin  is  nothing  but  these  Free  Receptors.— 
It  acts  like  a  lightning-conductor.  Withdrawn  from  the  body 
which  produces  it  and  introduced  into  another  animal,  it 
retains  the  same  fixing  property  and  is  the  active  agent  in 
therapeutic  sera. 

Thus  there  is  no  essential  difference  between  the  receptors 
which  produce  the  antitoxin  and  the  normal  "nutrition 


FIG.  70. — Diagrams  to  represent  Ehrlich's  theory. 

1.  Haptophore  group  h  and  toxophore  group  t. 

2.  Cell  C  injured  at  / :  its  receptor  r.     The  toxin  is  represented  by  T 
with  its  haptophore  group  h  and  its  toxophore  group  t. 

3.  A  toxin  molecule  :  haptophore  group  h  :    toxophore  group  t :  anti- 
body or  free  receptor  a.  r. 

4.  A  cell  C  with  a  fixed  receptor  r :  a  detached  free  receptor  forming 
the  antibody  a.  r.  i  the  toxin  T  with  its  haptophore  and  toxophore  groups 
h  and  t. 

receptors "  of  the  cell.  A  cell  susceptible  to  the  poison  pro- 
duces an  antidote,  but  the  antidote  may  also  be  produced  by 
cells  which  are  insusceptible,  /.*.,  not  only  by  the  "noble" 
cells,  but  also  by  the  connective-tissue  cells,  and  it  is  necessary 


IMMUNITY  215 

to  add,  in  the  light  of  the  experiments  of  the  Metchnikoff 
school,  by  the  leucocytes. 

In  bacteriolysis  and  haemolysis  as  defined  by  Bordet's 
experiments,  also  immunity  phenomena,  the  complement  and 
the  amboceptor  (or  immune-body)  come  into  play.  Com- 
plement is  an  atom  group  already  present  in  the  body,  whereas 
amboceptor  is  analogous  to  the  antitoxin :  it  possesses,  how- 
ever, two  haptophore  groups,  one  uniting  with  the  cells  (blood 
corpuscles  or  bacteria),  the  cytophil  group,  the  other  linking 
up  the  complement,  the  complementophil  group.  The  anti- 
bodies are  receptors  or  amboceptors  set  free  and  detached 
from  the  cells  which  form  them. 

It  is  impossible  to  follow  here  all  the  developments  of  the 
side-chain  theory.  Ehrlich  has  complicated  it  almost  to 
excess  in  order  to  include  in  it  the  infinite  complexity  of  facts 
observed  in  experiments  with  the  various  antibodies,  anti- 
toxins, haemolysins,  bacteriolysins,  and  the  other  cytolysins, 
precipitins,  and  agglutinins.  The  dominant  idea  is  always  to 
give  a  chemical  interpretation  of  nutrition  and,  as  a  particular 
case  of  this,  of  immunity. 

The  conception  was  first  intended  to  explain  the  physiology 
of  toxin  and  antitoxin  action,  and  it  is  to  this  that  it  is  best 
adapted ;  but  it  has  had  to  undergo  complication,  not  only  to 
include  the  largest  number  possible  of  the  ascertained  facts, 
but  to  act  up  to  its  original  intention  as  an  explanation  in 
terms  of  chemistry. 

It  is  to  explain  all  these  facts 1  that  the  distinction  between 

1  For  example  these,  that  the  broth  culture  of  the  diphtheria  bacillus 
which  constitutes  crude  toxin  is  not  a  simple  substance  :  nor  is  there  any 
reason  to  believe  that  the  broth  cultures  of  the  tetanus  bacillus  is  any 
more  so,  since  it  contains  at  least  two  poisons,  tetanolysin  and  tetano- 
spasmin.  The  products  of  cell-life  are  frequently  very  complex  :  Ehrlich, 
for  example,  quotes  with  justice  cinchona  bark  with  its  twenty  odd 
alkaloids  and  the  liver  cells  with  their  round  dozen  ferments. 
Toxin  left  to  itself,  even  protected  from  light  and  heat,  rapidly  becomes 
modified  :  it  feels  the  effect  of  age  and  not  only  deteriorates  in  activity  but 
undergoes  qualitative  changes.  Toxin  acts  after  a  period  of  incuba- 
tion. The  neutralization  by  antitoxin  no  longer  proceeds  in  the  same 
way  when  the  appropriate  dose  of  antitoxin  is  added  hi  separate  fractions 
instead  of  all  at  once  (Danysz-Dungern  phenomenon). 


216  MICROBES   AND  TOXINS 

the  haptophorous  and  toxophorous  groups  has  been  conceived, 
and  that  in  toxin  the  toxones  and  toxoids  have  been  postu- 
lated. But  the  complexity  is  also  due  to  the  necessity  of 
making  the  facts  conform  to  the  idea  of  the  chemical  nature 
of  these  actions,  and  especially  to  the  law  of  multiply  which 
demands  that  the  same  quantity  of  toxin  should  always  be 
neutralized  by  the  same  quantity  of  antitoxin.  The  theory 
flatters  itself  also  on  its  capacity  to  explain  the  specificity  of 
the  antibodies  ;  for  the  complexity  of  the  groups  is  supposed 
to  be  sufficiently  great  to  permit  a  haemolysin  against  goat 
corpuscles  to  differ  from  a  hsemolysin  against  rabbit  corpuscles. 
Even  for  the  fairly  numerous  cases  in  which  specificity  is  not 
rigorous  an  explanation  is  forthcoming:  different  receptors 
possess  certain  elements  in  common,  and  it  is  possible  that  a 
serum  which  precipitates  horse  serum  may  also  precipitate  the 
serum  of  the  ass. 

The  side-chain  theory  has  been  of  great  service ;  it  has,  its 
supporters  say,  a  great  "  heuristic  "  value,  /.£.,  it  has  been  the 
means  of  discovering  many  interesting  phenomena,  and  has  led 
Ehrlich  on  to  his  chemotherapeutic  studies,  in  which  he  has 
gained  such  magnificent  successes. 

Nevertheless,  it  cannot  be  said  that  these  fortunate  results 
prove  the  truth  of  the  theory :  the  discovery  of  "  606,"  for 
example,  proves  neither  the  existence  of  the  side-chains  nor  the 
truth  of  the  chemical  theory  of  immunity, 

Theory  of  Chemical  Equilibria.— Again  to  explain  the 
action  of  toxin  and  antitoxin,  Arrhenius,  Madsen  and  Walbum 
have  proposed  another  chemical  theory.  They  criticize  the 
complexity  of  the  theory  of  Ehrlich,  they  do  not  admit  the  com- 
plex nature  of  the  diphtheria  toxin,  and  they  attach  great  im- 
portance to  the  experiment  of  Danysz-Dungern  on  partial 
saturation.  The  facts  can  be  explained  by  conceiving  the 
toxin  +  antitoxin  reaction  as  a  chemical  interaction,  not 
between  a  strong  acid  and  a  strong  base,  but  between  a  weak 
acid  and  a  weak  base,  for  example,  ammonia  and  boric  acid. 
The  reaction  toxin  and  antitoxin  is  comparable  with  those 
reactions  known  as  reversible  and  is  governed  by  the  law  of 


IMMUNITY  217 

mass-action  of  Guldberg  and  Waage,  if  one  supposes  a  state 
of  unstable  equilibrium  to  exist  among  the  combinations.1 

The  combination  toxin  +  antitoxin  must  therefore  be  dis- 
sociable ;  Arrhenius  and  Madsen  are  of  the  opinion  that  they 
have  demonstrated  this  by  their  experiments  on  the  diffusion 
of  the  mixture  in  a  column  of  solidified  gelatine  and  by  their 
distribution  experiments  (e.g.,  distribution  of  agglutinin  between 
bacteria  and  the  immersing  fluid).  Many  facts  which  led 
Ehrlich  to  his  hypothesis  on  the  toxins  are  explained  by 
Arrhenius  in  terms  of  these  dissociations,  and  in  general  by 
the  fact  that  antibodies  and  antigens  have  only  a  feeble  affinity 
for  each  other. 

To  the  diffusion  experiments  it  has  been  urged  in  reply  that 
the  dissociation  only  takes  place  because  the  mixture  poured 
on  the  gelatin  has  not  had  time  to  form  the  final  combination ; 
while  to  the  hypothesis  that  a  quantity  of  free  toxin  is  present 
in  the  mixture,  it  has  been  replied  that  if  this  were  the  case 
antitoxin  would  never  act  in  the  body;  the  body  would  fix 
the  free  toxin,  the  toxin-antitoxin  equilibrium  would  be  dis- 
turbed, a  new  quantity  of  toxin  would  be  set  free,  and 
so  on. 

Physical  chemists  regard  the  dominant  idea  of  Arrhenius's 
theory  with  great  reserve ;  they  doubt  the  justice  of  employing 
the  laws  of  chemical  equilibrium  and  rates  of  reaction  in 
speculations  as  to  the  reactions  which  go  on  between  bodies  of 
which  nothing  is  known  from  the  chemical  point  of  view. 
Nernst  has  verified  the  use  made  of  the  laws  of  reversible 
actions,  and  he  denies  the  possibility  of  applying  to  colloidal 
substances  laws  established  only  for  homogeneous  liquids. 

This  does  not  mean  that  there  is  anything  odd  in  attempting 

1  When  a  substance  in  solution  of  a  molecular  concentration  m  reacts 
with  another  substance  in  solution  in  concentration  «,  the  mass  of  the 
substance  formed  by  their  combination  is  in  a  given  time  proportional  to 
the  product  mn. 

For  example,  when  in  a  given  volume  3  molecules  of  alcohol  react  with 
2  molecules  of  acetic  acid,  the  quantity  of  acetic  ether  formed  in  a  given 
time  is  3  x  2  =  6 ;  if  5  molecules  of  alcohol  react  with  3  molecules  of 
acetic  acid  the  quantity  of  acetic  ether  formed  in  the  same  time  is  expressed 


218  MICROBES  AND  TOXINS 

to  apply  physico-chemical  laws  to  biological  phenomena  in 
spite  of  the  variability  of  living  creatures  and  of  their  products. 
It  is  impossible  for  biologists  to  refrain  from  seeking  quanti- 
tative laws  and  from  applying  physico-chemical  laws  to 
immunity  phenomena.  Quantitative  results  have  been  ob- 
tained in  the  study  of  diastases,  and  Ehrlich  has  discovered 
facts  of  the  greatest  interest  by  means  of  his  experiments 
in  vitro  on  titrations  and  measuring.  It  is  only  necessary  to 
agree  upon  the  conventions  necessary  (in  physics  itself  these 
cannot  be  dispensed  with)  and  not  to  employ  such  unjustifiable 
expressions  as  " gukrison  in  vitro  "  (in  vitro  cure). 

It  is  always  possible  to  return  to  the  biological  point  of 
view  when  this  becomes  necessary,  as  Ehrlich  himself  did, 
when  considering  Weigert's  ideas  on  the  regeneration  of 
protoplasm. 

THE  PHYSICAL  POINT  OF  VIEW. 

Bordet  rejected  Ehrlich's  system,  and  compared  the  "  anti- 
gens +  anti-bodies  "  reactions  to  phenomena  of  absorption  and 
molecular  adhesion,  even  before  the  closer  comparison  with 
colloidal  reactions  had  been  arrived  at. 

He  does  not  only  complain  that  the  side-chain  theory  is  too 
complicated ;  he  criticises  its  whole  disposition. 

Immunity  is  a  problem  not  yet  ripe ;  and  the  solution  will 
probably  come  from  a  quarter  quite  unexpected.  There 
are  enormous  gaps  in  our  knowledge.  Why,  then,  make 
adventurous  generalizations  when  the  biological  facts  are  far 
from  permitting  this  ?  Every  theory  that  can  be  constructed 
must  base  itself  for  the  moment  on  facts  not  yet  demonstrated. 
Let  us  keep  close  to  the  experiments,  and  be  content  to 
advance  step  by  step.  Ehrlich's  theory  is  dangerous,  in  that 
it  offers  too  readily  conceptions  which  have  the  appearance  of 
explanations,  and  which,  therefore,  are  apt  to  dull  the  appetite 
for  research.  "  For  my  part,"  adds  Bordet,  "  I  have  been 
unwilling  to  construct  a  theory  ;  I  do  not  adduce  any  general 
conceptions ;  the  hypotheses  which  I  have  proposed  are 


IMMUNITY  219 

scarcely  worthy  of  the  name,  they  differ  so  little  from  mere 
critical  statements  of  experimental  results.  Even  at  the  risk 
of  being  regarded  as  incapable  of  generalizing,  I  prefer  to  stick 
to  the  facts  without  moulding  them  into  a  system." 

In  immunity  phenomena,  we  observe  certain  activities  ;  but 
why  materialize  them  and  picture  each  by  an  atomic  group  ? 
In  the  side-chain  theory  we  are  told  that  the  antibody  is 
nothing  but  a  cell-receptor  affected  by  the  antigen.  This 
identity  is  not  proved.  Why  should  not  the  cell  with  its 
power  of  adaptation  and  reaction  produce  some  new  and 
original  substance  ? 

When  the  "  substance "  which  we  call  agglutinin  clumps 
cells  or  bacteria,  does  it  really  bring  into  action  an  atomic 
group,  or  side-chain,  which  attaches  itself  and  another  group 
which  agglutinates  ?  The  explanation  is  artificial.  In  reality 
it  is  not  the  agglutinin  which  agglutinates  ;  it  is  a  salt.  The 
antigen  (bacteria)  and  the  antibody  (agglutinin)  form  a  com- 
bination which  produces  floccules,  or,  as  it  is  expressed 
nowadays,  is  "  flocculable  "  by  electrolytes.  It  is  this  couple 
or  "  complex  "  which  agglutinates.1 

An  analogous  coupling  must  be  regarded  as  taking  place  in 
all  the  reactions  of  antigens  and  antibodies,  and  Ehrlich's 
theory  is  wrong  in  attributing  everything  to  the  antibodies 
and  nothing  to  the  antigen.  There  are  no  "  amboceptors " 
in  reality,  there  only  exist  "  uniceptors "  capable  of  being 
absorbed. 

It  was  therefore  not  with  the  purpose  of  disputing  details 
that  Bordet  accumulated  his  experiments  on  the  mode  of 
fixation  of  the  complement  on  the  immune  body ;  in  this 
field  he  has  discovered  the  principal  facts  which  render  the 
side-chain  theory  impossible  as  a  dogma,  if  not  altogether 
impossible  as  a  conception  of  certain  phenomena.  The 
important  fact  is  that  there  is  never  absorption  of  complement 
by  an  immune  body  without  the  presence  of  an  antigen,  so 

1  The  salt  acts  on  bacteria  saturated  with  agglutinin  but  it  acts  also  on 
bacteria  which  have  absorbed  various  chemical  substances,  iron,  uranium, 
or  aluminium. 


220  MICROBES  AND  TOXINS 

true  is  it  that  it  is  the  antigen-antibody  compound  which 
absorbs  the  complement.  Although  the  complement  of  one 
animal  species  may  differ  from  that  of  another  species,  yet  in 
a  given  serum,  in  opposition  to  Ehrlich's  theory,  there  exists  but 
one  complement  or  rather  one  complementing  property  (exp. 
of  Border.,  Gay,  Muir  and  Browning,  etc.).  It  is  due  to 
Bordet's  correct  attitude  on  these  points  that  the  Bordet- 
Gengou  reaction  (complement  fixation)  has  been  capable  of 
such  successful  application  in  various  bacteriological  diagnostic 
methods,  and  recently  by  Wassermann  in  the  diagnosis  of 
syphilis. 

Finally,  since  the  complement  attaches  itself,  not  to  the 
immune  body,  but  to  the  antigen-antibody  combination,  there 
is  no  reason  to  suppose  the  existence  of  a  haptophore  group 
of  the  "  complementophile "  kind,  indispensable  to  the 
immune  body  if  complement  is  to  be  fixed.  This  question 
has  for  some  time  been  a  sort  of  test  for  the  side-chain 
theory,  and  it  seems  to  have  resulted  in  favour  of  Bordet's 
ideas.1 

From  the  beginning  of  his  researches  in  1896,  Bordet  has 
imagined  immunity  reactions,  not  as  chemical  combinations, 
but  as  physical  phenomena  of  absorption  or  molecular  adhesion. 
He  considered  that  in  agglutination  (where  the  bacteria  are 
passive,  since  dead  bacteria  also  agglutinate)  serum  acts  by 
modifying  the  relations  of  molecular  attraction  between  the 
bacteria  and  the  fluid  bathing  them,  and  that,  in  the  first 
phase  of  the  phenomenon  at  least,  the  bacteria  behave  like 
particles  in  general.  Under  the  influence  of  Duclaux's  ideas, 
he  observed  the  resemblances  between  agglutination  and 
coagulation.  From  the  point  of  view  of  their  coagulating  and 
dissolving  properties,  he  compared  the  active  sera  to  the 
digestive  juices,  and,  like  Metchnikoff  and  after  him  Ehrlich, 
he  also  saw  in  immunity,  though  from  a  different  point  of 
view,  a  particular  case  of  the  physiology  of  digestion. 

At    that   time   the   results   of    the   study   of   colloids   had 

1  Experiments  of  Ehrlich  and  Sachs,  of  Sachs  and  Bauer ;  of  Bordet 
and  Gay  and  Bordet  and  Streng  on  haemolysis  by  ox-serum. 


IMMUNITY 

scarcely  begun  to  be  applied  to  the  study  of  immunity,  and 
the  body-fluids,  the  toxins,  and  the  ajititoxins  had  not  yet 
been  studied  from  the  point  of  view  of  their  colloidal  con- 
stitution. The  phenomenon  of  adsorption  (ad  in  preference 
to  ab  as  expressing  the  idea  of  attraction  or  adhesion)  is  a  very 
general  one,  and  does  not  depend  absolutely  on  the  colloidal 
state.  Bordet  therefore,  in  explaining  his  conception,  pre- 
ferred to  employ  the  comparison  with  dyeing  processes.  The 
action  of  antitoxin  on  toxin  appeared  to  him  to  resemble,  for 
example,  the  action  of  iodine  on  starch  :  the  immune  body 
which  prepares  bacteria  or  cells  for  the  action  of  complement 
he  regarded  as  acting  after  the  manner  of  mordants  in  dyeing, 
intermediary  substances  necessary  for  the  fixing  of  certain  dyes 
on  certain  cloths.  Thus  in  haemolysis  the  union  of  the 
immune-body  with  the  blood  corpuscle  (anti-body  +  antigen) 
forms  a  combination  possessing  a  greater  adsorptive  affinity 
than  the  normal  corpuscle :  the  complement  tends  to  become 
precipitated  on  the  sensitized  corpuscle,  and  the  attraction 
which  the  latter  exerts  is  more  powerful  the  more  heavily  it  is 
sensitized.1 

Inorganic  substances  present  similar  phenomena.  Water 
runs  off  a  watch  glass  coated  with  paraffin  without  sticking  to 
it,  but  if  the  water  contains  barium  sulphate  in  suspension  it 
wets  the  paraffin  and  spreads  over  it ;  this  depends  on  the  fact 
that  the  surface  of  the  paraffin  becomes  coated  by  molecular 
adhesion  with  a  thin  white  film  of  barium  sulphate,  which 
water  can  wet ;  this  film  is  not  removed  by  rinsing  in  water, 
and  can  only  be  removed  by  rubbing.  There  are  even  sub- 
stances which  inhibit  this  fixation  of  the  barium  sulphate  on 
the  paraffin,  just  as  there  are  substances  which  inhibit  the 
fixation  of  complement  by  sensitized  corpuscles. 

On  all  the  most  important  points  of  the  question  of  the 
toxin  and  antitoxin  combination,  the  physical  theory  is  the 

1  Those  sera  which  possess  the  power  of  inhibiting  haemolysis  act  by 
keeping  the  complement  in  a  condition  of  greater  suspension  or  dissemina- 
tion in  the  fluid  ;  they  thus  render  it  more  stable,  unlike  saline  solution, 
which  produces  a  condition  of  instability  in  which  the  complement 
condenses  itself  or  is  precipitated  on  the  attracting  sensitized  cells. 


MICROBES  AND  TOXINS 

reverse  of  the  chemical  theory  of  Ehrlich ;  the  one  simplifies 
where  the  other  complipates. 

In  the  chemical  theory  the  same  quantity  of  antitoxin  ought 
to  combine  with  the  same  quantity  of  toxin,  so,  to  account  for 
the  irregularities  in  the  actual  facts,  there  have  been  introduced 
the  hypotheses  of  the  haptophore  group  separate  from  the 
toxophore,  of  the  toxones,  toxoids,  &c.  On  the  contrary, 
Bordet  supposes  that  the  antitoxin  really  unites  with  the  toxin 
in  varying  proportions  :  the  toxin  can  fix,  can,  so  to  speak, 
dye  itself  with  antitoxin  in  greater  or  less  amount  just  as  starch 
can  take  up  varying  quantities  of  iodine  and  become  thereby 
stained  a  more  or  less  dark  blue.  In  the  same  way,  in 
haemolysis  the  corpuscles  can  absorb  variable  amounts  of  the 
active  substance  according  to  the  concentration  of  the  solutions 
and  the  duration  of  the  contact.  The  distance  of  this  idea 
from  the  theory  of  chemical  equivalents  is  apparent.  When  a 
given  quantity  of  toxin  is  mixed  with  a  quantity  of  antitoxin 
insufficient  for  complete  neutralization,  what  occurs  is  not  a 
monopolization  of  the  antitoxin  by  a  portion  of  the  toxin  mole- 
cules, forming  a  complete  combination  with  it  while  the  rest  of 
the  toxin  remains  free  (i.e.,  the  chemical  conception).  On  the 
contrary,  the  antitoxin  is  equally  distributed  over  all  the  toxin 
present,  so  that  the  latter  is  attenuated  throughout  and  presents 
a  diminished  activity.  To  return  to  the  same  analogy,  it  is 
faintly  dyed.  It  can  produce  toxic  effects  qualitatively  different 
from  those  due  to  an  intact  toxin  or  a  toxin  completely 
neutralized  without  necessitating  the  hypothesis  of  a  special 
chemical  condition  (toxones).1 

The  phenomenon  of  Danysz-Dungern  (the  antitoxin  has  a 
different  action  on  toxin  when  the  mixture  of  the  same 
quantities  is  made  at  one  instead  of  several  additions)  does  not 
compel  the  hypothesis  that  in  toxin  there  are  several  chemically 

1  The  fact  first  observed  by  Ehrlich,  namely,  the  difficulty  of  preparing 
exactly  neutral  mixtures  of  toxin  and  antitoxin,  can  thus  be  easily  explained. 
The  effects  produced  by  a  hsemolysin  more  or  less  neutralized  and  the 
experiments  of  Grossberger  and  Schattenfroth  on  the  toxin  and  antitoxin  of 
the  bacillus  of  quarter-evil  have  confirmed  Bordet's  views  of  the  nature  of 
the  toxin  +  antitoxin  reaction. 


IMMUNITY 

distinct  components.  If  a  large  piece  of  filter-paper  is  dipped  in 
a  rather  dilute  solution  of  a  dye  it  is  faintly  stained;  if  the 
piece  is  cut  into  small  pieces  and  these  are  immersed  in  turn 
for  a  certain  time,  we  find  that  the  first  pieces  take  up  the  colour 
and  leave  almost  none  for  the  last.  Substitute  for  the  dye  the 
antitoxin  and  for  the  paper  the  toxin  ;  if  the  mixture  is  made  at 
one  blow  the  toxin  is  attenuated  in  its  whole  bulk  :  if,  however, 
the  mixture  is  made  by  several  additions,  the  first  portions  of 
the  toxin  take  up  the  antitoxin  and  the  later  portions,  not  being 
neutralized,  remain  much  more  toxic. 

Further,  the  fact  that  toxin  and  antitoxin  mixtures  (and  in 
general  antigens  and  antibodies)  become  in  time  less  dissolvable 
and  more  stable  is  also  explained  by  adsorption  phenomena. 

When  a  piece  of  cloth  is  placed  in  a  dyeing  vat  the  dye 
loses  its  attachment  to  the  dissolving  fluid  and  adheres  more 
and  more  intimately  to  the  cloth  until  it  can  no  longer  be 
redissolved.  A  similar  example  may  be  quoted  in  connection 
with  the  precipitates  produced  by  alcohol  in  certain  albuminous 
fluids,  precipitates  which  are  fairly  easily  redissolved  in  water 
immediately  after  their  precipitation,  but  which  are  no  longer 
soluble  when  a  certain  time  has  been  allowed  to  elapse  for 
their  aggregation.  -When  three  substances  exist  together,  two 
of  them  may  compete  with  each  other  in  the  fixation  of  the 
third  ;  it  is  thus  that  the  protective  action  of  certain  substances 
is  to  be  explained,  as,  for  example,  the  albuminous  substances 
of  the  blood  which  protect  blood  corpuscles  against  the  action 
of  soap  (Meyer)  or  of  eel-serum  (Frouin).1 

One  thing  that  the  physical  theory  has  not  yet  explained  is 
the  specificity  of  the  reactions  of  immunity,  but  it  is  not 
incapable  of  explaining  even  this. 

It  is  not  difficult  to  imagine  that  slight  physical  modifica- 
tions may  change  the  affinities  on  which  depend  the  molecular 
attraction ;  that  is  certainly  no  more  difficult  to  imagine  than 

1  Citrate  of  soda  protects  blood  corpuscles  against  the  agglutinating  and 
haemolytic  action  of  sulphate  of  barium.  The  lecithin  of  ox-seram  is  held 
in  check  as  regards  its  action  on  guinea-pigs'  corpuscles  by  some  albuminoid 
material. 


MICROBES   AND  TOXINS 

the  innumerable  molecular  groups  postulated  by  the  side-chain 
theory.  For  example,  the  antiserum  prepared  by  an  animal 
against  a  protein  has  not  the  same  properties  as  the  serum 
obtained  against  the  same  substance  previously  subjected  to 
heat  (exp.  of  Obermayer  and  Pick).  Hens'  serum  agglutinates 
the  lipoids  extracted  from  the  red  corpuscles  of  the  rabbit 
much  more  vigorously  than  those  from  the  ox.  It  is  easy  to 
found  a  theory  for  specificity  on  the  absorption  phenomena, 
and  such  theories  already  exist ;  hitherto  they  have  been  too 
philosophical,  but  it  is  satisfactory  to  know  that  experiment 
has  already  furnished  the  germ  of  a  scientific  explanation. 

The  Colloids. — After  Bordet's  explanations  of  agglutination 
and  haemolysis  in  terms  of  molecular  attractions  and  cohesions, 
and  in  the  light  of  his  comparison  of  these  phenomena  to 
dyeing  processes,  Zangger,  Landsteiner,  and  Jagic  established 
experimentally  the  first  analogies  between  immunity  phenomena 
and  the  physics  of  colloids. 

Reactions  between  colloids  or  between  colloids  and  true 
solutions  can  be  reduced  to  phenomena  of  molecular  attrac- 
tion, of  absorption,  and  adsorption.  The  bodies  participate 
in  the  reactions  in  variable  proportions,  influenced  by  tem- 
perature and  pressure.  Colloids  of  opposite  electrical  charge 
exert  on  each  other  a  "flocking"  or  precipitating  action, 
which  may  be  masked  when  one  or  other  is  in  excess  in  the 
mixture ;  one  colloid  may  inhibit  the  precipitation  of  another 
colloid  by  a  salt.  In  many  cases,  the  law  of  the  opposite 
electrical  charge  may  be  masked  by  the  fact  that  the  proteins 
are  amphoteric  colloids,  and  may  neutralize  acids  and  alkalies 
equally  well,  behaving  in  acid  solution  as  bases,  in  basic 
solutions  as  acids. 

Are  the  antigens  and  antibodies  of  immunity  colloids  ? 
The  only  ones  whose  chemical  composition  is  known,  namely, 
the  lipoids  (fatty  bodies  typified  by  lecithin  or  cholesterin), 
behave,  in  watery  suspension,  like  the  colloids  ;  the  others, 
which  probably  belong  to  the  proteins,  behave  like  colloids 
from  the  point  of  view  of  diffusion  in  dialysis,  heat,  and 
instability,  and  their  principal  reactions  are  closely  analogous 


IMMUNITY  225 

to  those  of  colloids.  In  any  case,  there  might  exist  between 
the  antibodies  and  antigens  on  the  one  hand,  and  the  colloids 
on  the  other,  considerable  differences  without  preventing  the 
general  laws  of  attraction  and  adsorption  from  being  equally 
applicable  to  the  former  as  to  the  latter. 

Agglutination  and  precipitation  closely  resemble  the  flocking 
of  colloids.  Bacteria  behave  towards  a  precipitating  serum  in 
the  same  way  as  they  behave  to  certain  substances  quite 
foreign  to  the  body,  such  as  gelatine  or  gum-arabic. 

Bacteria,  in  presence  of  a  solution  of  ferric  chloride,  are 
protected  by  the  colloidal  ferric  hydrate  from  agglutination  by 
their  specific  agglutinin. 

The  phenomena  of  specific  haemolysis  (i.e.,  haemolysis  by 
the  serum  of  immunized  animals)  have  been  imitated 
by  attacking  the  corpuscles  by  means  of  such  systems  as 
silicic  acid  +  lecithin,  colloidal  ferric  hydrate  +  dog- 
serum,  or  saponin  +  taurocholate  of  sodium  :  the  two  kinds 
of  phenomena  may  be  expressed  by  the  same  curves 
(Zangger,  Mile.  Cernovodeanu,  and  V.  Henri).  Comple- 
ment can  be  fixed  by  the  most  diverse  substances,  by 
Witte  peptone,  yeast  cells,  the  cells  of  organs,  and  various 
precipitates. 

The  toxin  +  antitoxin  reaction  has  been  "  imitated  "  by  the 
interaction  of  arsenious  acid  and  colloidal  ferric  hydrate.  The 
phenomenon  of  Danysz-Dungern  can  also  be  reproduced  with 
colloids ;  the  final  result  in  the  precipitation  of  a  colloidal 
suspension  is  different  according  as  the  precipitant  is  added  in 
one  large  or  in  fractional  doses. 

The  reactions  between  cholesterin  and  various  poisons  and 
between  cholesterin  and  lecithin  have  the  appearance  of 
colloidal  reactions.  One  may  even  observe  affinities  of  the 
character  of  the  specific  affinities.  For  example,  cholesterin 
neutralizes  saponin  and  tetanolysin,  but  has  no  effect  on  ricin 
or  staphylolysin.  Complement  is  absorbed,  as  has  been  seen, 
by  a  great  variety  of  substances  ;  agglutinin,  however,  possesses 
affinity  chiefly  for  the  colloidal  protein,  while  tetanus  toxin  is 
chiefly  fixed  by  the  lipoids.  It  is  legitimate  to  conceive  of 

Q 


226  MICROBES   AND   TOXINS 

these  affinities  as  depending  upon  physical  conditions,  including 
that  of  the  electrical  charges. 

The  lipoid  substances  which  are  found  in  such  abundance  in 
nerve-tissue  and  which  constitute  a  constant  component  of 
protoplasm,  according  to  Overton  furnish  the  cell  with  a  sort 
of  envelope  through  which  the  food  materials  have  to  pass  and 
which  behaves  as  a  sort  of  colloidal  atmosphere ;  the  principal 
members  of  this  group  are  lecithin  and  cholesterin.  It  is 
inconceivable  that  they  do  not  play  some  part  in  immunity 
phenomena  which  are  phenomena  of  nutrition.  Haemolysin, 
for  example,  undoubtedly  induces  changes  in  the  lipoid  coating 
of  the  red  blood  corpuscles. 

The  lipoid  extracts  of  red  corpuscles  readily  fix  normal 
hsemolysins,  whereas  the  lipoid  extracts  of  bacteria  fix  certain 
immune-bodies.  The  lipoids  also  are  capable  of  fixing  the 
complements. 

We  have  already  mentioned  two  series  of  experiments  in 
which  lipoids  play  a  most  definite  part.  In  the  first  place  we 
have  the  activation  of  cobra  venom  by  lecithin  (Kyes),  in  which 
the  latter  appears  to  play  the  part  of  complement.  According 
to  Noguchi,  triolein,  oleic  acid,  exerts  the  same  effect  and  loses 
its  activity  quite  like  complement  when  heated.  Oleic  acid  is 
said  to  be  capable  of  activating  specific  hsemolysins,  and  silicic 
acid,  which  alone  possesses  a  very  feeble  hsemolytic  power,  is 
said  to  form  with  lecithin  a  complex  or  "  lecithid "  which  is 
much  more  powerful. 

In  the  second  place  lecithin  plays  the  part  of  antitoxin 
towards  certain  toxins.  The  bile  or  the  soluble  elements 
of  bile  neutralize  snake  venom  in  appropriate  dose ;  the 
cholesterin  is  the  active  constituent.  Cholesterin  and  lecithin 
neutralize  the  botulismus  toxin.  Cholesterin  neutralizes 
saponin,  solanin,  agaricin,  vibriolysin,  lecithids  of  cobra  venom 
and  of  the  poison  of  bees :  again  it  is  its  cholesterin  content 
which  makes  serum  neutralize  saponin.  Finally  in  the 
celebrated  experiment  of  the  fixation  and  neutralization  of 
tetanus  toxin  by  the  cerebral  cortex  (Wassermann  and  Takaki's 
experiment)  the  lipoids  of  the  grey  matter  are  the  active 


IMMUNITY 

agents ;  the  brain  extracted  with  ether  loses  much  of  its 
neutralizing  power,  and  the  dye  carmine  neutralizes  because  it 
contains  lipoids  derived  from  the  cochineal  insect  (Metchnikoff). 

It  is  facts  such  as  these  which  encourage  investigators  to 
pursue  the  line  opened  up  by  Bordet's  experiments,  although 
there  is  no  absolute  promise  that  they  will  find  in  this  the  key 
to  immunity. 

If  we  add  now  that  phagocytosis  is  not  incompatible  either 
with  Bordet's  theory  or  with  Ehrlich's,  it  is  true  enough,  but  it 
is  not  all  the  truth.  It  is  not  a  question  of  reconciling 
theories.  There  are  only  two  "  theories,"  that  of  Ehrlich  and 
that  of  Bordet,  which,  with  their  conjectures,  their  uncertainty, 
their  attempts  at  explanation,  and  their  continual  state  of 
incompleteness,  are  striving  to  round  off  the  positive  doctrine, 
the  expression  of  undoubted  facts,  namely  phagocytosis. 
When  a  physiologist  is  studying  digestion  he  founds  his  study 
on  facts  which  are  essential  and  certain,  such  as  the  action  of 
trypsin  and  enterokinase  ;  this  is  no  theory  ;  it  is  only  when 
he  proceeds  to  interpret  this  action  in  terms  of  physics  or 
chemistry  by  fixations  or  combinations  that  he  enters  the 
domain  of  theory.  Similarly,  it  is  no  way  of  recognizing  the 
capital  importance  of  phagocytosis  to  admit  that  antibodies 
and  other  humoral  properties  are  produced  by  the  phagocytes. 
The  essential  fact  is  the  destruction  of  the  microbes  by 
incorporation  and  digestion.  Extraphagocytic  destruction  is 
so  much  an  exceptional  case  that  it  cannot  even  be  brought  in 
as  opposition. 

There  would  be  no  temptation  to  forget  this  fact  if,  instead 
of  limiting  our  attention  to  human  pathology,  we  kept  in  view 
the  universality  of  intracellular  digestion  throughout  the  series 
of  living  beings.  Phagocytosis  is  quite  different  from  a  medical 
theory.  It  is  a  doctrine  as  fundamental  in  general  biology  as 
is  the  existence  of  the  cell  or  the  variation  of  species. 

Toxins  and  Phagocytosis. — The  immunity  of  an  animal 
towards  a  toxin  is  not  to  be  ascribed  simply  to  the  activity  of 
its  body-fluid.  We  must  take  account  of  the  body  also  in  the 
reaction.  There  are  many  facts  indicating  a  similar  state  of 

Q  2 


228  MICROBES   AND  TOXINS 

affairs,  as  in  the  experiment  of  Heymans  and  Masoin  on  the 
neutralization  in  vivo  of  hydrocyanic  acid  by  hyposulphite  of 
soda. 

In  vivo  the  hyposulphite  of  soda  acts  as  an  antidote  or 
chemical  antitoxin  to  hydrocyanic  acid.  Now  no  one  has  ever 
succeeded  in  reproducing  this  experiment  in  the  test-tube, 
whereas  in  the  body  it  is  perfectly  easy.  "  In  consequence  it 
is  legitimate  to  appeal  to  certain  peculiar  conditions  in  the 
living  animal,  which,  however,  does  not  exclude  the  possibility 
that  the  transformation  of  the  toxic  substance  into  a  harmless 
material  may  be  due  to  a  chemical  reaction." 

Fresh  nutrient  broth  possesses  an  antitoxic  action  towards 
abrin  intoxication  (Calmette).  The  serum  of  an  animal 
immunized  against  certain  toxins  or  venoms  protects  other 
animals  to  a  greater  or  less  extent  against  the  action  of  other 
toxins  or  other  venoms ;  here  there  can  be  no  question  of  a 
specific  direct  antitoxic  action. 

The  fresh  blood  of  the  crayfish  is  capable  of  preventing  the 
fatal  intoxication  of  mice  by  scorpion  venom ;  yet  the  crayfish 
is  killed  by  a  dose  of  scorpion  venom  three  or  four  times 
smaller  than  that  necessary  to  kill  a  mouse,  and  the  blood  of 
the  crayfish  has  no  protective  action  for  another  crayfish. 

Roux  and  Vaillard  observed  long  ago  that  animals  might  die 
of  tetanus  although  possessing  an  abundance  of  antitoxin  in 
their  blood.  There  are  certain  horses  originally  good  furnishers 
of  diphtheria  or  tetanus  antitoxin  which  suddenly  cease  to 
produce  this  in  their  serum  although  they  remain  immune. 

Rabbits  may  be  immunized  against  tetanus  by  inoculating 
them  under  the  skin  of  the  tail  several  times  in  succession 
with  tetanus  spores  along  with  a  little  lactic  acid ;  the  animal 
becomes  resistant  to  the  toxin  and  yet  100  volumes  of  its 
serum  fail  to  neutralize  a  single  minimum  lethal  dose  of 
toxin  (Vaillard).  The  antitoxic  power  of  the  body-fluids  is 
thus  not  sufficient  to  explain  acquired  immunity,  since  it  is 
not  an  invariable  fact  in  animals  rendered  immune. 

The  actions  of  the  body  itself  have  again  to  be  reckoned 
with  in  attacking  the  question  of  the  origins  of  antitoxins. 


IMMUNITY  229 

We  have  already  mentioned  the  old  opinions  of  Buchner  and 
Roux.  Buchner  considered  that  the  antitoxin  was  derived 
from  the  toxin,  and  Metchnikoff  advanced  the  opinion  that 
certain  body-cells  might  produce  this  transformation.  But, 
it  has  been  protested,  how  could  a  horse  react  to  a  single  unit 
of  toxin  by  producing  100,000  units  of  antitoxin  ?  The  toxin 
may,  however,  be  seized  by  certain  organs  which  retain  it  for 
a  long  period  and  transform  it  slowly.  The  toxin  may  induce 
in  the  cells  which  produce  antitoxin  the  very  stimulus  which 
Ehrlich  was  among  the  first  to  appeal  to.  The  experiment  of 
Roux  and  Vaillard  on  rabbits  immunized  against  tetanus 
would  thus  be  explained :  after  repeated  bleedings  the 
antitoxin  power  of  the  blood  rapidly  regained  its  former 
titre.  But  why  should  the  serum  of  healthy  animals  sometimes 
have  a  certain  antitoxic  power?  Because  without  having 
actually  suffered  from  diphtheria  or  tetanus  they  may  have 
harboured  diphtheria  or  tetanus  bacilli  in  their  bodies  :  in 
the  intestine  of  the  horse  for  example  the  tetanus  bacillus 
abounds. 

Whether  or  not  the  toxin  is  transformed  into  antitoxin,  it  is 
certain  that  the  antitoxin  is  a  product  of  the  body  :  no  other 
way  of  producing  it  is  known.  Ehrlich  formerly  thought  that 
the  cells  sensitive  to  the  toxin  were  its  chief  producers ;  but 
if  this  were  true,  antitoxin  ought  to  be  present  in  these  cells 
and  be  capable  of  neutralizing  the  toxin.  Wassermann  and 
Takaki's  experiment  seemed  to  prove  this  :  the  brain  tissue  of 
mammals  ground  up  with  tetanus  toxin,  neutralizes  it,  furnish- 
ing a  mixture  which  no  longer  gives  tetanus  to  animals.  But 
Wassermann's  experiment  has  in  reality  a  quite  different 
signification.  The  brain  does  not  act  as  an  antitoxin,  for, 
if  injected  into  a  guinea-pig  along  with  a  dose  of  toxin,  but  at 
separate  points  in  the  body,  it  has  no  protective  action  what- 
ever, and  does  not  act  in  the  least  like  a  dose  of  antitoxin. 
Further,  if  the  so-called  neutral  mixture  is  injected  into  the 
thigh  of  a  guinea-pig,  the  animal  becomes  tetanic,  whereas  it 
remains  protected  if  the  injection  is  made  into  the  peritoneum. 
The  neutralizing  property  is  in  reality  a  property  peculiar  to 


230  MICROBES   AND  TOXINS 

the  cerebral  cortex  (and  the  grey  matter  of  the  spinal  cord)  of 
mammals  only ;  in  fowls  immunised  against  tetanus  the  brain 
has  much  less  neutralising  power  than  the  blood,  the  liver,  or 
the  kidney.  The  brain  of  the  frog  does  not  neutralise  the 
toxin,  although  under  certain  conditions  the  frog  is  susceptible 
to  tetanus. 

Cholesterin,  lecithin,  and  even  olive  oil  and  carmine  (a 
substance  derived  from  the  fatty  body  of  the  cochineal  insect) 
are  capable  of  neutralizing  a  certain  quantity  of  toxin ;  now 
the  brain  substance  contains  both  cholesterin  and  lecithin. 
It  is,  however,  another  lipoid  in  the  brain,  protagon,  which 
chiefly  fixes  the  toxin  and  perhaps  permits  of  its  transport 
along  the  nerves  (Landsteiner  and  Botteri.)  A.  Marie  and 
Tiffeneau  have  recently  insisted  that  in  Wassermann's  experi- 
ment it  is  not  a  destruction  which  takes  place,  but  a  combina- 
tion from  which  the  toxin  may  be  recovered.  Anyhow,  the 
neutralization  by  the  cerebral  tissue  is  a  phenomenon  of 
molecular  adhesion,  analogous  to  a  dyeing  process. 

By  injecting  the  tetanus  toxin  directly  into  the  brain  it  has 
been  shown  that  this  very  brain  substance  which,  ground  up 
in  a  glass,  neutralizes  the  toxin,  does  not  neutralize  in  the 
living  animal  the  most  minute  dose.  It  is  therefore  impossible 
to  suppose  that  the  brain  is  a  source  of  antitoxin  (Roux  and 
Borrel). 

An  immunized  rabbit,  rendered  resistant  to  toxin  by  subcu- 
taneous inoculation,  succumbs  when  the  toxin  is  injected  into 
the  brain  :  the  antitoxic  action  is  therefore  due  to  cells  lying 
between  the  periphery  and  the  centre ;  the  poison  is  neutralized 
en  route. 

What  are  these  cells?  We  find  that  sublethal  doses  of 
tetanus  toxin  produce  in  the  fowl  a  great  afflux  of  leucocytes 
into  the  blood ;  further,  in  a  fowl  injected  with  tetanus  toxin, 
far  less  toxin  is  to  be  found  in  the  blood  than  in  (aseptic) 
exudates  rich  in  leucocytes.  Metchnikoff  therefore  considers 
that  the  protection  of  the  body  against  toxins  also  depends  on 
the  leucocytes. 

The  rabbit  can  stand  large  doses  of  atropin  injected  subcu- 


IMMUNITY  231 

taneously  or  into  the  circulation,  but  it  is  very  susceptible 
to  intracerebral  injection ;  is  it  the  leucocytes  which  dispose 
of  the  poison  when  injected  into  the  veins  ?  This  is  possible, 
for  the  poison  can  be  recovered  from  the  leucocytes  after  it 
has  disappeared  or  is  only  to  be  found  in  traces  in  the  blood 
plasma  (Calmette). 

When  guinea-pigs  are  injected  intraperitoneally  with  arsenic 
trisulphide,  a  salt  which  is  only  slightly  soluble,  the  particles 
are  taken  up  by  the  macrophages.  When  a  soluble  salt  like 
potassium  arsenite  is  injected,  when  the  animal  recovers  it  is 
in  the  leucocytes  that  the  arsenic  is  found  on  chemical  analysis 
(Besredka).  The  phagocytes  also  absorb  lead  salts  (Carles). 
The  white  corpuscles  of  the  blood  are  thus  equally  capable  of 
resisting  mineral  as  microbial  poisons. 

When  a  guinea-pig  is  inoculated  with  the  mixture  of 
mammalian  brain  ground  up  with  tetanus  toxin  the  solid 
particles  attract  the  phagocytes  which  seize  them  and  with 
them  the  attached  toxin.  If  the  toxin  is  injected  along  with 
particles  to  which  it  does  not  attach  itself  (e.g.  the  mixture  of 
toxin  +  frog's  brain)  it  can  diffuse,  and  the  leucocytes  no 
longer  protect  the  animal. 

After  the  discovery  of  the  antitoxins  one  was  apt  to  think 
that  in  every  infection  there  was  an  intoxication  and  that  the 
neutralization  of  the  poisons  ought  to  precede  phagocytosis 
which  is  only  a  secondary  phenomenon.  But  experience  has 
shown  that  the  phagocytes  can  also  digest  the  microbial  toxins. 

Certain  bacteria  secrete  substances  which  weaken  and 
destroy  the  phagocytes  ;  among  these  antiphagocytic  microbial 
poisons  may  be  classed  the  agresstns  of  Bail.  Now  the 
phagocytes  are  capable  of  absorbing  and  digesting  these 
substances  without  any  outside  assistance. 

Certain  bacterial  extracts  prepared  outside  the  body  and 
injected  in  sufficient  quantity  are  prejudicial  to  phagocytosis. 
Yet  the  same  bacteria  which  furnish  these  extracts  are 
absorbed  by  the  leucocytes  when  these  latter  have  their 
activity  reinforced. 

When  dead  typhoid  bacilli  are  injected  into  the  peritoneum 


MICROBES   AND   TOXINS 

of  a  guinea-pig  they  are  of  course  incapable  of  producing  an 
infection,  but  as  they  contain  the  typhoid  endotoxin  the 
animal  dies  nevertheless  from  the  intoxication.  But  if  before 
the  injection  the  peritoneum  is  "prepared"  so  that  the 
bacteria  at  once  meet  with  a  great  number  of  vigorous  leuco- 
cytes the  poison  is  absorbed  by  these  and  the  animal  is  saved. 

A  staphylococcus  habituated  to  the  rabbit  by  the  method  of 
passages,  secretes  a  poison  which  is  very  injurious  to  the 
leucocytes  of  this  animal ;  but  if  along  with  the  staphylococci 
vigorous  living  leucocytes  are  injected  into  the  pleura,  the 
rabbit  is  protected  against  the  intoxication  (Bail  and  Weil). 

Immunity  against  toxin  therefore  can  be  reduced,  like  the 
immunity  against  bacteria,  to  the  simple  fact  of  phagocytic 
digestion.  Metchnikoff  finds  himself  justified  in  the  light  of 
all  these  experiments,  in  thinking  that  it  is  the  phagocytes 
which  take  up  poisons  and  which  perhaps  themselves  elaborate 
the  antitoxins  employed  in  serotherapy. 


CHAPTER  XII 

ANAPHYLAXIS 

Definition  of  anaphylaxis— Experiments  of  Richet  and  Portier — Anaphylaxis 
to  various  poisons — Anaphylaxis  to  normal  serum — Arthus'  phenomenon 
—Serum  sickness  :  observations  of  V.  Pirquet  and  Schick — Serum 
anaphylaxis  in  guinea-pigs  ;  phenomenon  of  Th.  Smith — Anaphylaxis 
to  cells  and  organ  extracts — Passive  anaphylaxis. 

Study  of  Richet's  poisons  and  serum  anaphylaxis— Anti-anaphylaxis 
(Besredka)  :  not  a  vaccination — Application  to  serotherapy — Heating 
of  sera — Theories  of  anaphylaxis — General  theory  of  antibodies. 

ANAPHYLAXIS  is  the  opposite  of  vaccination.  An  animal  is 
vaccinated  when  the  first  attack  by  a  virus  (microbe  or  toxin) 
produces  in  it  changes  through  which  it  is  protected  against 
another  more  serious  attack.  If  you  suppose  the  animal  to 
become,  after  the  first  attack,  not  more  resistant,  but  more 
susceptible,  you  have  in  a  word  the  root  idea  of  anaphylaxis. 

The  paradox  is  even  greater  than  this  definition  shows,  for 
anaphylaxis  exists  not  only  towards  viruses  against  which 
immunity  also  occurs,  but  also  it  appears  towards  substances 
which  in  the  normal  individual  are  apparently  quite  innocuous, 
for  example,  egg-white,  milk,  and  serum.  In  this  way  the 
first  injection  with  these  substances,  instead  of  producing 
immunity,  seems  to  destroy  the  natural  immunity  of  the  normal 
animal. 

The  supersensitiveness  produced  by  a  first  inoculation  and 
brought  to  light  by  a  second  is  not  due  to  a  cumulative  action, 
such  as  may  be  observed  after  several  successive  doses  of 
certain  drugs ;  in  anaphylaxis  the  effect  is  out  of  all  proportion 
to  the  quantity  of  material  ingested. 


234  MICROBES   AND  TOXINS 

This  chapter  of  experimental  medicine  was  opened  up  by 
the  experiments  of  Richet  and  Portier  on  the  poison  of 
Actinians  (1902). 

Anaphylaxis  to  Poisons— Richet's  Experiments.— 
From  the  tentacles  of  actinians  a  poison  may  be  extracted 
which  has  been  called  congestin,  because  it  produces  in  the 
animals  injected  an  intense  congestion  of  the  viscera,  of  the 
stomach,  liver,  kidney,  and,  above  all,  the  intestine.  The 
fatal  phenomena  do  not  appear  until  after  several  hours  of 
incubation.  If  a  dog  which  has  recovered  from  a  small  dose  is 
inoculated  after  a  certain  interval  of  time  with  -fa  of  the 
quantity  of  the  first  dose,  violent  symptoms  suddenly  appear 
after  a  few  seconds,  severe  vomiting,  difficult  respiration, 
paralysis,  profuse  and  blood-stained  diarrhoea.  On  comparing 
the  size  of  the  dose  with  its  effect,  it  works  out  that  the 
dog  has  become,  between  the  first  injection  and  the  second, 
eighty  times  more  susceptible. 

Anaphylaxis  to  Normal  Serum — Arthus'  Phe- 
nomenon.— If  a  rabbit  is  injected  subcutaneously  every  six 
days  with  5  c.c.  of  horse  serum,  it  is  found  that  after  the  fifth 
injection  the  serum  is  absorbed  with  difficulty ;  succeeding 
injections  produce  local  lesions  which  continually  increase  in 
severity,  from  simple  inflammation  to  actual  necrosis  of  the 
tissue.  The  same  phenomenon  may  be  produced  with  milk 
instead  of  serum. 

"  Serum-sickness  "  :  Observations  of  Von  Pirquet 
and  Schick. — Therapeutic  sera  (antidiphtheria,  antitetanus) 
are  almost  always  got  from  immunized  horses ;  they  are  not 
invariably  devoid  of  toxicity  for  man.  In  a  small  proportion 
of  cases,  about  14  per  100,  the  injection  is  followed  by  various 
symptoms,  not  very  severe  it  is  true,  which  only  appear  after 
an  incubation  period  of  five  to  fifteen  days,  and  consist  of 
urticaria,  erythemas,  and  pains  in  the  joints.  But  should  the 
patient  fall  ill  again  months  or  years  later,  long  after  the  serum 
has  disappeared  from  his  body,  and  should  he  be  reinjected 
with  it,  the  symptoms  are  reproduced  and  are  more  frequent 
(86  per  100,  according  to  Weil-Halle  and  Lemaire),  more 


ANAPHYLAXIS  235 

intense,  and  more  rapid ;  they  appear  within  an  hour,  or  even 
within  a  quarter  of  an  hour,  after  the  injection. 

In  the  light  of  these  observations  V.  Pirquet  observed  later 
that  the  unknown  germs  of  vaccine  lymph  produce  a  premature 
reaction  in  the  skin  of  those  individuals  formerly  vaccinated 
and  now  supersensitive.  The  same  observation  in  its  turn 
suggested  to  him  the  idea  of  applying  a  droplet  of  tuberculin 
to  the  skin  of  a  tuberculous  patient  to  test  his  susceptibility, 
and  this  was  the  origin  of  the  cutaneous  reaction  of 
tuberculosis. 

Through  these  observations  the  study  of  anaphylaxis  entered 
the  sphere  of  human  medicine,  and  the  important  question 
arose,  How  are  we  to  render  harmless  our  therapeutic  sera  ? 

Serum  Anaphylaxis  in  the  Guinea-pig.  The 
Phenomenon  of  Th.  Smith. — The  question  entered  the 
laboratories  when  attempts  were  made  to  elucidate  the  fact 
observed  in  the  American  serotherapeutic  institutes,  that 
guinea-pigs  which  had  already  been  employed  in  the  titration 
of  antidiphtheritic  serum  (injected  with  a  mixture  of  toxin  and 
serum)  became  eventually  supersensitive  to  horse  serum. 
If  three  to  twelve  weeks  after  the  titration  injection  5  c.c.  of 
horse  serum  is  injected  under  the  skin  or  especially  into  the 
peritoneum,  the  animals  immediately  manifest  anxiety  and 
discomfort ;  respiration  is  rapid  and  laboured,  the  heart 
becomes  weaker,  the  temperature  falls  below  normal  and  after 
one  hour  50  per  cent,  of  the  animals  die,  whereas  normal 
guinea-pigs  support  doses  of  the  same  serum  five  times  as  great 
without  any  disturbance.  The  phenomenon  is  specific,  for 
guinea-pigs  receiving  horse  serum  in  the  first  dose  behave 
on  re-injection  quite  like  normal  guinea-pigs  towards  rabbit, 
goat  or  ox  serum.  In  the  toxin  +  antitoxin  titration  mixture 
the  antitoxin,  i.e.,  the  horse  serum,  is  responsible  for  the 
supersensitiveness  induced.  A  small  dose  sensitizes  more 
certainly  than  a  large  dose;  even  one  millionth  of  a  c.c. 
may  suffice  !  The  symptoms  resemble  serum-sickness  in  man ; 
there  is  an  incubation  period  of  twelve  days  at  least  after  the 
first  injection.  The  supersensitive  condition  persists  for 


236  MICROBES   AND   TOXINS 

months  (experiments  of  Otto,  Rosenau  and  Anderson, 
Besredka). 

Anaphylaxis  to  Cells. — Animals  injected  with  red 
corpuscles,  washed  (to  prevent  the  action  of  the  serum)  or  un- 
washed, resist  the  first  injection  well  but  support  a  second 
badly. 

Similarly  with  bacterial  cells,  typhoid,  paratyphoid  bacilli, 
etc.,  they  present  more  or  less  severe  symptoms  after  the 
second  injection.  The  specificity  of  this  reaction  does  not 
appear  to  be  very  strict.  This  variety  of  anaphylaxis  is  of 
great  practical  importance  for  in  the  preparation  of  anti- 
plague  sera  it  is  necessary  to  inject  the  cultures  into  the 
jugular  vein  of  horses,  and  severe  symptoms  are  quite 
common. 

Organ  Extracts. — The  phenomena  induced  by  injections 
of  extracts  of  spleen,  lymphatic  glands,  bone  marrow,  or 
spermatozoa  are  analogous  to  the  preceding.  The  supersensi- 
tive  state  can  be  induced  towards  extracts  of  the  crystalline  lens 
of  the  eye  and  an  animal  thus  prepared  reacts  only  towards  the 
lens  tissue,  no  matter  from  what  species  of  animal,  even  when 
it  presents  no  reaction  to  the  serum  of  that  animal. 

A  tuberculous  patient  becomes  supersensitive  to  the  product 
of  the  tubercle  bacillus,  to  tuberculin.  But  in  this  case  it 
appears  that  only  the  disease  itself  can  induce  this  excessive 
susceptibility  permanently  :  it  is  very  doubtful  if  it  can  be 
produced  by  the  injection  of  the  bacterial  bodies  or  of 
tuberculin.  Physiologically  speaking  it  is  an  anaphylactic 
phenomenon. 

Passive  Anaphylaxis.— M.  Nicolle  has  shown  that  if  a 
fresh  rabbit  is  given  a  large  dose  of  the  serum  of  a  rabbit 
rendered  anaphylactic  to  horse  serum,  the  fresh  rabbit  takes  on 
an  anaphylaxis  which  is  therefore  called  passive.  It  may  in  its 
turn  present  the  typical  symptoms  on  injection  24  hours  after  the 
preparatory  injection.  This  brief  delay  shows  that  it  is  a  true 
passive  anaphylaxis,  and  not  an  active  anaphylaxis  induced  by 
small  quantities  of  horse  serum  which  might  have  remained  in 
the  serum  of  the  sensitive  rabbit.  This  experiment  demon- 


ANAPHYLAXIS  237 

strates  the  existence  of  an  antibody  in  the  serum  of  the  sensitive 
animal ;  this  antibody  has  been  transferred  to  the  other  animal 
just  as  the  diphtheria  antitoxin  of  the  horse  can  transfer  passive 
immunity  to  man. 

Passive  anaphylaxis  can  be  produced  particularly  well  by 
injecting  the  serum  of  an  anaphy lactic  rabbit  into  a  normal 
guinea-pig.  Passive  transference  has  also  been  produced  in  the 
case  of  Richet's  poisons.  It  is  therefore  a  general  fact,  although 
experiments  of  transference  between  animals  of  the  same  species 
do  not  always  succeed. 

From  these  fundamental  experiments  there  have  proceeded 
several  sets  of  researches  ;  especially  with  the  poisons  and  with 
serum  considerable  advances  have  been  made.  It  is  not  at  all 
certain  in  spite  of  their  manifest  similarities  that  the  laws  of 
anaphylaxis  against  poisons  and  serum  (or  other  non-toxic  pro- 
tein) are  really  entirely  the  same.  There  is  always  this  differ- 
ence, that  in  the  one  case  it  is  a  question  of  substances 
manifestly  toxic  to  the  normal  body,  whereas  in  the  other  the 
substances  are  such  that  the  healthy  animal  shows  no  visible 
reaction. 

For  the  practical  point  of  view,  these  researches,  both 
on  the  poisons  and  on  the  sera  (because  of  serotherapy),  are 
of  obvious  value;  there  is  an  actual  disease  to  prevent  or 
cure. 

It  does  not  seem  as  if  the  anaphylaxis  to  egg-white  should 
interest  us  to  the  same  extent.  We  do  not  take  albumen  or 
milk  by  subcutaneous,  intraperitoneal,  or  intravenous  injections. 
If,  as  we  know  already,  our  body  tends  to  resist  the  introduc- 
tion of  foreign  albumens,  we  have  a  digestive  tube  which  trans- 
forms these  into  our  own  specific  protein ;  that  is  perhaps  why 
so  few  examples  exist  hitherto  of  anaphylaxis  acquired  by  taking 
food.  Further,  it  is  probable  that  we  are  much  more  exposed 
to  the  cumulative  action  of  toxic  bodies,  e.g.,  the  phenols  of  the 
digestive  tube,  than  to  an  anaphylaxis  to  the  proteins  of  the  ox 
or  of  the  fowl.  But  the  digestive  tube  may  occasionally  be 
defective  as  a  defensive  agent. 

Poisons  also  may  be  absorbed  by  other  routes,  such  as  the 


238  MICROBES   AND  TOXINS 

pulmonary  alveoli  or  the  skin.  We  are  here  in  a  region  as  yet 
little  explored ;  there  are  many  mysteries  in  the  action  of  drugs, 
there  are  still  greater  mysteries  in  the  experimental  study  of 
diseases  of  nutrition.  Once  again  anaphylaxis  raises  this 
general  problem  of  nutrition,  towards  which  already  the  whole 
study  of  immunity  is  directed,  and  it  is  probable  that,  did  we 
know  more  of  this,  we  would  be  less  ignorant  of  the  nature  of 
life,  old  age,  and  death. 

Let  us  consider  then  separately  what  knowledge  has  been 
gained  on  the  subject  of  anaphylaxis  to  those  substances  which 
have  been  most  studied,  the  poisons  and  serum. 

Researches  on  the  Poisons. — There  have  been  studied 
by  Richet  the  congestin  of  Actinians,  the  congestin  extracted 
from  mussels,  and  a  vegetable  toxin,  analogous  to  abrin  and 
ricin  (well-known  from  Ehrlich's  experiments),  called  crepitin, 
and  extracted  from  the  plant  Hura  crepitans  of  the  Euphor- 
biaceae,  known  in  Brazil  under  the  name  of  Assaku.  The 
mytilocongestin  (from  mussels)  produces  vomiting,  a  symptom 
very  definite  and  easy  to  observe,  a  great  convenience  in 
experimental  work.  "The  symptoms  of  anaphylaxis  to 
crepitin  are  exactly  the  same  as  with  actino-  or  mytilo-con- 
gestin,  and  even  the  most  acute  observer,  I  am  sure,  could  not 
distinguish  them.  .  .  .  There  is  the  same  profound  abolition 
of  all  innervation,  both  motor  and  sensory,  and  above  all, 
vaso-constrictory :  there  is  the  same  intense  hsemorrhagic 
congestion  of  the  intestine  with  an  enormous  fall  in  the 
arterial  pressure  "  (Richet). 

It  is  to  be  observed  that  these  belong  to  the  group  of  slow 
poisons  —  resembling  thus  the  bacterial  toxins — which  differ 
from  the  crystalloid  group  such  as  strychnine,  and  it  is  probable 
that  on  this  depends  their  anaphylactic  effect. 

An  animal  is  rendered  anaphylactic  because  the  first 
injection  has  induced  in  it,  after  a  period  of  incubation,  the 
formation  of  a  new  substance,  an  antibody,  the  product  of  a 
reaction  of  the  body.  This  antibody  is  not  itself  poisonous, 
but  it  liberates  a  poison  when  it  comes  in  contact  with  the 
congestin  or  crepitin  of  the  second  injection. 


ANAPHYLAXIS  239 

The  antibody  is  named  by  Richet  toxogeninc,  and  the  new 
poison,  apotoxine.  He  therefore  imagines  anaphylaxis  after  the 
formula :  Toxogenine  +  Congestine  =  Apotoxine. 

He  considers  that  the  apotoxin  results  from  the  action  of 
the  two  substances  on  each  other,  the  one,  congestin,  introduced 
from  without  and  non-toxic  in  the  dose  employed  for  the  test 
injection,  the  other,  toxogenin,  non-toxic,  prepared  by  the 
body  itself  as  a  result  of  the  first  small  injection  of  congestin. 
In  a  similar  way  we  have  emulsin  and  amygdalin  acting 
together  to  produce  hydrocyanic  acid. 

At  first  the  existence  of  this  toxogenin  was  hypothetical,  but 
experiment  has  in  two  ways  proved  its  real  existence — i.  When 
a  normal  animal  is  injected  with  the  blood  of  a  sensitized 
animal,  it  becomes  anaphylactic  without  any  period  of  incubation, 
i.e.,  there  is  passive  anaphylaxis.  2.  Richet  has  several  times 
succeeded  in  demonstrating  the  reaction  with  crepitin  in  vitro. 
The  mixture  in  a  test-tube  of  anaphylactic  serum  and  poison, 
when  injected  after  incubation,  produces  immediate  anaphylactic 
effects  :  the  test-tube  contains  both  the  antibody  and  the 
antigen.  3.  The  toxogenin  does  not  exist  only  in  the  blood ;  it 
is  also  to  be  found  in  the  brain  tissue.  By  mixing  crepitin  with 
brain  substance  (freed  from  blood)  or  even  with  the  alcoholic 
precipitate  from  the  brain  substance  of  an  anaphylactic  animal, 
immediate  anaphylactic  symptoms  can  be  produced  in  a  fresh 
animal :  there  is  thus  "  cerebral  anaphylaxis  "  in  vitro.  There  is 
thus  antibody  present  in  the  nerve-cell  and  capable  of  reacting 
with  the  antigen  at  the  moment  of  the  anaphylactic  shock. 
It  is  this  very  reaction  which  Besredka  regards  as  the 
essence  of  the  anaphylactic  shock,  which  appears  to  him  to  be 
eminently  a  cerebral  phemonenon. 

According  to  Richet  those  animals  which  have  received 
an  anaphylactising  toxin  become  from  this  sole  fact  more 
sensitive  to  other  poisons.  The  injection  of  an  antigen,  for 
example  crepitin,  renders  an  animal  more  sensitive  to  toxic 
action  of  other  kinds,  for  example  to  that  of  apomorphine, 
although  the  increase  of  sensitiveness  is  particularly  directed 
towards  the  antigen  itself.  There  exists  therefore,  it  would 


240  MICROBES   AND  TOXINS 

seem,  a  sort  of  general  anaphylaxis  in  addition  to  the 
specific  form  of  anaphylaxis.  Richet  is  inclined  to  consider 
that  in  the  phenomena  of  anaphylaxis  there  is  in  reality  only 
a  relative  specificity,  and  that  the  apotoxin  is  a  poison  without 
any  specificity  which  attacks  and  paralyses  the  central  nervous 
system,  affecting  in  this  particularly  the  vasomotor  centres. 
"  It  seems  to  me  very  probable  that  in  the  study  of  the  whole 
field  of  the  different  forms  of  anaphylaxis  produced  by  different 
substances  a  great  analogy  if  not  identity  would  be  discovered 
in  the  symptoms  of  all  forms  of  anaphylaxis,  so  that  we  may  be 
permitted  to  believe  in  the  general  analogy  if  not  in  the  identity 
of  all  the  different  apotoxins,  the  various  anaphylactic  poisons. 
There  would  thus  be  a  very  simple  conclusion,  namely,  that 
there  is  one  poison  and  one  only,  the  apotoxin  produced  in 
all  the  forms  of  anaphylaxis." 

Richet's  poisons  produce  in  the  body,  besides  anaphylaxis, 
immunity.  With  mytilocongestin  the  anaphylaxis  disappears 
after  about  six  weeks  and  the  immunity  persists.  Ana- 
phylaxis thus  gives  place  to  its  opposite  "prophylaxis"  and 
toxogenin  to  antitoxin.  The  two  conditions  "  develop  side  by 
side  from  the  moment  of  the  first  injection.  Hence  it  is 
necessary  to  distinguish  closely  between  the  immediate  and  the 
late  effects.  During  the  anaphylactic  period  there  is  a  striking 
supersensitiveness  as  regards  the  immediate  symptoms,  but 
there  is  already  some  immunity  towards  the  late  effects  of  the 
poison.  If  the  animal  survives  the  immediate  effects  of  the 
second  dose,  it  presents  no  further  symptoms  during  the  days 
following.1.  .  .  Anaphylaxis  is  the  first  stage  in  prophylaxis" 
These  views  recall  those  of  Behring.  "  However  para- 
doxical it  may  seem,  there  can  be  no  doubt  that  horses  which 
have  become  strongly  immune  as  the  result  of  treatment  with 
tetanus  toxin  yet  possess  a  histogenic  supersensitiveness  of 
their  tissues  towards  the  tetanus  toxin."  These  properties  are 

1  Richet  applies  the  same  idea  to  serum -anaphylaxis  and  to  anaphylaxis 
in  general.  "  The  anaphylactic  reaction  is  a  defensive  function  and  is 
destined  to  maintain  intact  the  chemical  constitution  and  homology  of 
each  animal  species  by  preventing  foreign  albumins  from  entering  the 
protoplasm  of  the  cells  so  as  to  modify  their  specific  chemical  structure." 


ANAPHYLAXIS  241 

probably  common  to  many  toxins,  but  in  serum  anaphylaxis 
the  same  characteristics  do  not  necessarily  exist. 

Anaphylaxis  to  poisons  thus  means  a  hastening  of  the 
reaction  of  the  body  towards  microbial  toxins  :  it  is  a  process 
adapted  for  rapid  defence  and  in  particular  for  defence  against 
small  doses  :  it  is  a  sort  of  alarm  signal  sent  to  the  body-cells 
calling  its  attention  to  minute  quantities  of  poison  which 
without  this  would  have  been  insufficient  to  induce  immunity  : 
immunity  is  established  thanks  to  the  preceding  anaphylaxis. 
Perhaps  anaphylaxis  is  equivalent  to  Ehrlich's  superexcitation 
of  the  cells  required  for  the  production  of  antitoxins. 

Serum  Anaphylaxis. — A  preparatory  injection  of  serum 
in  very  small  dose  (1/250  to  1/1,000,000  c.c.),  a  period  of 
incubation,  a  test  injection  larger  than  the  first  and  producing 
violent  symptoms,  these  are  the  conditions  of  serum 
anaphylaxis  in  the  guinea-pig. 

On  observing  the  symptoms  it  is  impossible  to  avoid  the 
idea  that  the  guinea-pig  has  fallen  victim  to  some  cerebral 
lesion  which  was  latent  during  the  period  of  incubation  but 
has  been  revived  by  the  test  injection.  The  symptoms  of 
this  lesion  ought  to  be  still  more  definite  if  the  injection 
were  made  into  the  brain  itself.  The  nerve-centres  can  be 
more  directly  attacked  by  intracerebral  or  intradural  in- 
jections or  through  the  circulation  by  the  intravenous  route 
(Besredka). 

One  quarter  of  a  c.c.  injected  intracerebrally  produces  the 
same  phenomena  as  5  c.c.  intraperitoneally  and  produces  them 
with  much  greater  regularity,  for  instead  of  about  25  per  cent., 
practically  all  the  guinea-pigs  die.  The  experiments  are  thus 
on  a  surer  footing.  Should  the  still  unknown  reaction  take 
place  in  the  brain  cell  itself,  i.e.t  the  reaction  between  the 
poison  and  the  antibody  manufactured  by  the  body  as  a 
result  of  the  first  injection,  one  might  hope  to  modify  the 
anaphylactic  phenomena  by  acting  on  the  nerve-cells  of  the 
supersensitive  animal.  And  this  is  really  the  case,  as  is  shown 
by  a  very  pretty  experiment  of  Roux  and  Besredka.  The 
sensitive  guinea-pig  is  put  to  sleep  with  ether  or  put  under  the 

m. 


MICROBES   AND  TOXINS 

influence  of  a  suitable  dose  of  alcohol ;  an  intracerebral 
injection  which  kills  the  sensitive  control  leaves  the 
narcotised  guinea-pig  unharmed.1 

In  practice  intracerebral  injections  have  supplied  Besredka 
with  a  useful  means  of  measuring  the  toxicity  of  a  serum,  for 
example,  anti-diphtheria  serum,  from  the  point  of  view  of  the 
symptoms  of  serum  sickness.  Should  a  method  be  discovered 
of  diminishing  or  destroying  the  toxicity  of  sera,  the  improve- 
ment acquired  could  thus  be  estimated.  In  particular,  the 
method  will  tell  us  whether  a  serum  may  be  safely  injected  by 
the  route  by  which  it  manifests  its  greatest  toxicity,  but  also  its 
greatest  efficacy,  i.e.,  intravenously.  Experience  has  shown 
that  a  serum,  very  toxic  the  day  of  the  bleeding  (minimum 
lethal  dose  ^  c.c.),  rapidly  loses  this  toxicity  during  the  ten 
days  following  (lethal  dose  ^  c.c.).  It  continues  to  diminish 
slowly,  for  a  month  and  a  half,  but 'after  about  two  months  it 
remains  almost  indefinitely  at  the  same  level  (lethal  dose  |  c.c.), 
and  never  becomes  non-toxic.  For  example,  a  sample  of 
serum  kept  in  Roux's  laboratory  for  thirteen  years  still  kills 
sensitive  guinea-pigs  in  a  dose  of  \  c.c.  intracerebrally 
(Besredka). 

Antianaphylaxis. — Anaphylaxis  is  a  morbid  state  predis- 
posing to  the  occurrence  of  accidents.  But  these  may  be 
prevented.  Antianaphylaxis  is  the  name  given  by  Besredka  to 
the  condition  of  insensibility  to  which  the  sensitive  animal  can 
rapidly  be  brought.  But  though  in  current  language  one  talks 

1  There  exist  other  physiological  shocks  which  are  deadened  by  narcosis, 
and  it  is  impossible  to  avoid  a  comparison  between  the  above  experiment 
and  one  of  Jellinek's  (the  director  of  the  laboratory  of  electrical  pathology 
in  Vienna)  reported  in  a  recent  lecture  by  D'Arsonval. 

"A  rabbit  is  fairly  easily  killed  when  the  opposite  poles  of  an  alternat- 
ing current  of  1,500  volts  are  placed  in  its  mouth  and  rectum,  whereas  a 
rabbit  of  the  same  breed  but  so  deeply  chloroformed  that  all  its  vital 
phenomena  have  ceased  is  at  once  reawakened  and  saved  from  death  by 
the  same  current. 

ic  At  the  time  that  this  experiment  was  published  an  English  engineer, 
Aspinall,  observed  that  two  electrical  engineers  who  had  during  sleep  come 
into  accidental  contact  with  an  alternating  current  of  3,000  volts  were 
simply  awakened  by  burning  sensations  in  the  back  without  other  injury." 
(Jellinek). 


ANAPHYLAXIS  243 

vaguely  of  "  vaccinating  "  against  anaphylaxis  by  Besredka's 
method,  antianaphylaxis  is  in  reality  not  a  vaccination. 

Originally  it  was  conceived  as  a  true  vaccination,  and  to 
produce  it  Rosenau  and  Anderson  made  a  series  of  injections 
of  5  c.c.  of  serum  intraperitoneally,  starting  before  the  period 
of  incubation  for  the  anaphylactic  state  was  up  :  they  proceeded 
as  one  does  for  antitoxins. 

But  this  idea  had  to  be  given  up  when  it  was  seen  that  it 
was  sufficient  to  protect  the  guinea-pig  to  give  it,  not  a  series  of 
injections,  but  a  single  injection,  not  a  large  injection,  but  a 
minimal  one,  -^  to  -^  c.c.,  *.*.,  much  less  than  the  toxic 
dose ;  finally,  and  above  all,  that  the  resistance  of  the  guinea- 
pig  develops  the  day  after,  or  even  some  hours  or  minutes 
after,  the  injection,  according  to  the  route  employed.  After 
subcutaneous  injection  the  resistance  is  present  in  four  or  five 
hours  ;  after  intradural  injection  in  about  two  hours ;  after 
intraperitoneal  in  an  hour  and  a  half;  ten  to  fifteen  minutes 
after  intravenous  injection.  The  shortness  of  this  latter  incu- 
bation period  may  be  employed  to  render  the  resistance  more 
and  more  complete  :  it  is  simply  a  question  of  repeating  these 
minute  doses  intravenously  every  ten  or  fifteen  minutes  :  each 
injection  reinforces  the  effect  of  the  preceding  :  this  is  the 
method  of  "  continuous  vaccination  "  (vaccinations  subintr antes). 
It  is  not  even  necessary  to  inject  by  the  veins :  by  taking  a 
little  more  time  one  may  proceed  by  other  routes  and  even 
changing  routes  in  successive  doses.  In  this  way  a  guinea-pig 
may  be  protected  from  as  many  lethal  doses  of  serum  as  is 
desired  (Besredka). 

In  this  there  is  the  germ  of  a  method  which  may  be  applied 
to  man.  Certain  sera,  in  particular  the  anti-plague  serum, 
have  to  be  injected  in  large  quantities,  and  as  far  as  possible 
by  the  veins.  The  anti-diphtheria  serum  is  500  times  more 
active,  other  things  being  equal,  if  introduced  directly  into  the 
circulation  instead  of  under  the  skin  (Berghaus).  Besredka's 
process  promises  to  relieve  such  intensive  serumtherapy  from 
all  danger  of  death  by  anaphylaxis. 

What  sort  of  immunity  then  is  this  which  develops  with  such 

R  2 


244  MICROBES   AND   TOXINS 

small  doses  and  so  rapidly  ? l  A  vaccinated  animal  departs  from 
the  normal  to  acquire  a  new  condition ;  in  sensitized  animals 
under  these  circumstances  there  is  not  the  departure  from  a 
normal  state  but  there  is  an  appearance  of  returning  towards  it. 
Antianaphylactic  vaccination  proceeds  like  a  disintoxication 
and  antianaphylatic  immunity  seems  to  be  a  return  to  the 
natural  immunity. 

The  sensitizing  injection  induces  the  formation  of  an  antibody ; 
at  the  moment  of  the  test  injection  it  is  probable  that  the  toxic 
serum  combines  with  or  rather  fixes  itself  on  this  antibody 
and  this  abrupt  reaction  produces  a  shock  which  the  nerve-cell 
resents  profoundly.  It  is  "  disintoxicated  "  by  it,  but  with  such 
brutal  suddenness  as  to  kill  the  animal.  When  by  narcosis  with 
anaesthetics  or  alcohol  the  sensitiveness  of  the  nerve-cell  is 
diminished,  as  in  the  experiment  of  Besredka  and  Roux,  it 
becomes  disintoxicated  during  sleep,  just  as  in  a  surgical 
operation  under  chloroform,  and  wakes  up  free.  If  instead  of 
injecting  as  in  the  test  injection  a  massive  toxic  dose,  a 
minute  dose  is  introduced  (^\^  c.c.  intravenously,  for  example), 
the  disintoxication  of  the  body,  and  in  particular  of  the  nerve- 
cells,  is  carried  out  gradually,  little  by  little  ;  in  the  continuous 
vaccination  method  the  disintoxication  leads  back  the  animal 
by  degrees  to  its  normal  condition. 

To  the  normal  condition,  but  not  quite,  for  the  injection 
leaves  as  a  rule  a  trace  of  serum  in  the  body  with  which  it 
re-sensitizes  itself  in  time ;  the  guinea-pig  has  become  a  fresh 
animal,  and  like  a  fresh  animal  may  be  sensitized.  It  may  lose 
its  virginity,  regain  it,  and  again  lose  it.  Certain  experimenters 
do  not  believe  that  the  body  ever  becomes  again  normal. 

The  digestive  tube  is,  physiologically  speaking,  as  much  a 
barrier  to^  the  invasion  of  the  body  by  foreign  proteins  as  it  is 

1  "  The  immunity  to  a  toxin  does  not  appear  till  after  eight  days  at 
least  and  is  more  pronounced  the  more  numerous  and  prolonged  the 
successive  doses  :  it  is  accompanied  by  the  appearance  of  the  antibody  in 
the  serum,  and  finally  it  never  extends  to  the  nerve-centres.  On  the 
contrary  anaphylactic  immunity  is  established  after  a  single  injection,  is 
practically  instantaneous,  is  never  accompanied  by  the  appearance  of 
antibodies  and  finally  always  extends  to  the  nerve-centres,  the  brain  and 
spinal  cord  "  (Besredka). 


ANAPHYLAXIS  245 

towards  bacteria  :  by  its  very  power  of  digesting  and  assimilat- 
ing them  it  renders  inoffensive  those  substances  which,  if 
injected  under  the  skin  or  into  the  blood  stream,  would  perhaps 
be  toxic  :  it  maintains  the  natural  immunity. 

But  for  the  present  it  must  be  pointed  out  that  Besredka's 
conception  only  applies  to  substances  like  serum  which  are 
harmless  in  general  to  the  normal  body  even  when  injected 
direct  into  the  veins.  It  is  not  necessarily  true  of  the  tox- 
albumins,  which  are  toxic  from  the  first. 

We  have  briefly  indicated  two  procedures  for  rendering  a 
sensitive  animal  resistant,  narcosis  during  the  test  injection  and 
the  method  of  continuous  vaccination  with  small  doses.  The 
same  result  may  be  obtained  by  heating  the  toxic  serum  to 
certain  definite  temperatures.  Heating  produces  an  effect  un- 
realized hitherto  by  any  chemical  means.  Serum  heated  to 
1 00°  C.  (diluted  to  prevent  coagulation)  becomes  practically 
harmless  for  both  intracerebral  and  intravenous  injections : 
But  this  heated  serum,  no  longer  lethal,  can  still  protect 
against  the  anaphylactic  shock  by  gradually  disintoxicating  the 
sensitive  cells  as  in  the  method  of  small  doses  or  narcosis ;  no 
doubt  the  mechanism  is  the  same. 

Heating  is  thus  a  good  means  of  lowering  the  toxicity  of  a 
serum.  But  sera  heated  to  100°  C.  lose  all  preventive  and 
curative  power,  and  it  is  impossible  therefore  to  exceed  59-60°  C. 
Experiment  shows  that  heating  for  several  hours  at  this  tem- 
perature diminishes  considerably  the  anaphylactic  toxicity. 
The  reason  for  the  low  toxicity  of  the  Pasteur  Institute  sera, 
duly  recorded  by  various  laboratories,  is  simply  that  their 
sterilization  is  effected  by  means  of  several  heatings  at  a  low 
temperature  (without  antiseptics). 

Man  is  not  alone  in  profiting  by  laboratory  research  on 
anaphylaxis.  In  certain  countries  the  Pasteur  vaccine  against 
anthrax  is  injected  along  with  a  few  c.c.  of  an  anti-anthrax 
serum :  the  method  is  named  sero-vaccination.  Many  cattle 
are  inoculated  over  again  every  two  years,  so  that  severe 
anaphylactic  effects  are  not  uncommon.  Recently  sero- 
vaccination  was  repeated  on  180  cattle  in  Roumania.  The 


246  MICROBES   AND   TOXINS 

animals  were  divided  into  two  lots,  the  first  receiving  i  c.c.,  as 
an  anti-anaphylactic  injection,  five  hours  before  the  sero- 
vaccination.  In  the  result  it  was  found  that  during  the  twenty- 
four  hours  following  the  sero-vaccination,  not  one  of  these 
showed  any  anaphylactic  symptoms,  whereas  ten  of  the  other 
ninety  showed  symptoms,  such  as  oedema  of  the  muzzle  with 
salivation,  oedema  of  the  vulvar  and  anal  mucous  membranes, 
and  colic  (Alexandresco  and  Ciuca). 

Theories. — No  final  theory  for  anaphylaxis  yet  exists,  but 
there  are  various  hypotheses  Awhich  keep  experimental  work 
active.  There  is  one  point,  however,  which  is  certain,  that  it 
is  impossible  to  explain  the  phenomena  without  the  presence  of 
an  antibody  formed  by  the  animal  as  a  consequence  of  the 
first  injection.  This  has  been  demonstrated  by  M.  Nicolle  in 
connection  with  Arthus's  phenomenon. 

It  is  probable  that  the  anaphylactic  shock  is  due  to  the  union 
of  antibody  and  antigen,  that  this  union  is  abrupt  and  affects 
especially  the  nerve-cells,  which  does  not  mean  that  the  nerve- 
cells  produce  the  antibody.  On  the  contrary,  everything  points 
to  their  not  creating  the  supersensitiveness,  but  being  passively 
affected  in  it. 

According  to  Vaughan  and  Wheeler,  who  studied  in  par- 
ticular the  anaphylaxis  to  egg-white,  the  sensitizing  injection,  as 
an  antibody,  provokes  the  formation  of  a.  preferment  which  can 
only  act  after  the  reinjection  by  splitting  the  protein  molecule 
into  two  components,  one  toxic,  the  other  not.  As  a  matter  of 
fact  it  is  possible  in  the  laboratory  to  produce  from  albumin 
two  such  components,  but  the  toxic  one  is  also  toxic  for  normal 
animals  quite  as  much  as  for  sensitive  ones :  this  therefore  is 
not  an  explanation  of  anaphylaxis,  though  it  shows  that  there 
exist  toxic  elements  in  substances  normally  non-toxic. 

It  was  natural  to  attempt  to  identify  the  antibody  of  anaphylaxis 
with  some  antibody  already  known  in  the  reactions  of  immunity. 
This  was  Friedberger's  idea  :  in  serum  anaphylaxis  the  antibody 
is  nothing  but  the  precipitin,  in  anaphylaxis  to  blood  corpus- 
cles a  haemolysin.  The  facts,  however,  do  not  confirm  Fried- 
berger's theory. 


ANAPHYLAXIS  247 

Besredka's  theory  approximates  more  closely.  In  anaphyl- 
axis  three  activities  come  into  play,  the  sensibilisinogen,  the 
property  of  serum  by  virtue  of  which  it  sensitizes,  the 
sensibilisin,  the  property  due  to  the  body  and  corresponding 
to  the  antibody  generally  admitted,  and  the  antisensibilisin,  by 
which  is  meant  the  property  of  normal  serum  in  virtue  of 
which  it  combines  with  the  sensibilisin  and  determines  the 
anaphylactic  shock. 

Thus  serum  is  toxic,  not  because  it  contains  a  poison  ready 
made,  but  because  two  substances,  non-toxic  themselves,  "  enter 
into  abrupt  combination  within  the  nerve-cells  and  thus 
disturb  their  equilibrium." 

Why  should  small  doses  sensitize  so  well,  whereas  massive 
doses  only  do  so  after  a  long  delay  ?  The  answer  is  that  a 
minute  dose  induces  the  formation  of  sensibilisin  without 
furnishing  it  with  anything  on  which  to  fix  :  in  consequence  it 
remains  avid,  in  the  nerve-cells  among  others.  After  a  large 
dose  the  sensibilisin  manufactured  is  neutralized  as  it  forms 
by  the  "  anti-sensibilisin  "  of  the  serum  and  anaphylaxis  is 
therefore  delayed.  It  seemed  probable  at  first  that  by  heating 
the  serum  at  suitable  temperatures  it  might  be  possible  to 
dissociate  the  two  activities  which  do  not  depend  on  the 
body,  i.e.,  the  sensitizing  and  toxic  properties.  But  on 
closer  examination  it  was  found  that  heating  acted  equally  on 
both  ;  the  two  in  reality  follow  the  same  curve.  It  had  also 
to  be  recognized  that  the  toxic  function,  the  sensitizing 
function,  and  the  vaccinating  or  anti-anaphlyactic  function,  are 
all  fundamentally  identical  and  occur  in  the  same  serum,  but 
that  they  correspond  to  different  physical  conditions  of  that 
serum. 

To  sensitize,  a  serum  must  be  injected  highly  diluted ;  to 
produce  the  anaphylactic  shock  it  must  possess  its  normal 
concentration ;  to  "  vaccinate  "  or  protect  against  the  shock  it 
must  unite  very  slowly  with  the  sensibilisin  of  the  body  or 
must  unite  with  it  in  minute  quantities  at  a  time  (e.g.,  in  the 
method  of  "  continuous  vaccination  "). 

Heated  at    100°  C  it  is   no   longer   toxic  or  very  slightly 


248  MICROBES   AND  TOXINS 

so,  because  being  partly  coagulated  its  fixation  is  delayed. 
The  action  of  serum,  sensitizing,  vaccinating,  or  toxic,  depends 
thus  on  its  physical  condition. 

There  are  therefore  really  only  two  activities  of  serum 
in  question,  that  of  the  antigen  and  that  of  the  antibody.  But 
according  to  its  physical  condition,  according  to  the  manner 
and  time  of  its  injection,  the  antigen  plays  the  part  of  sensitizer, 
of  vaccinator  or  of  toxin.  These  explanations,  it  must  be 
mentioned,  are  confined  to  the  field  of  serum-anaphylaxis  as  it 
is  known  to-day. 

We  always  return  thus  to  the  antigen  and  the  antibody,  i.e.,  to 
the  activities  which  we  found  to  exist  in  all  the  phenomena  of 
immunity. 

General  Theory  of  the  Antibodies — M.  Nicolle  has 
proposed  a  general  explanation  embracing  anaphylaxis  as  a 
particular  case  of  the  general  physiology  of  the  antibodies. 
Every  antigen  induces  in  the  body  the  simultaneous  formation 
of  antibodies  of  two  classes,  the  coagulins  and  the  lysins.  The 
coagulins  condense  albuminoids  and  toxins  (to  speak  only 
of  these  two  varieties  of  antigen) ;  the  lysins,  on  the  contrary, 
break  them  up  and  liberate  from  them  their  real  toxic 
components  : l  in  intoxication  by  proteid  poisons  it  is  not  these 
themselves  which  injure  the  body,  but  secondary  poisons 
elaborated  by  it  itself.  The  fate  of  the  animal  depends  on  its 
species,  on  the  nature  and  quantity  of  the  antigen,  on  the 
channel  of  inoculation  and  on  the  route  by  which  the  assaulting 
dose  is  introduced. 

Thus,  in  the  phenomenon  of  Th.  Smith,  the  supersensitive- 
ness  is  explained  by  the  development  of  a  lysin  and  the 
absence  of  all  coagulin  makes  of  it  the  type  example  of  pure 
supersensitiveness.  In  bacterial  anaphylaxis,  the  super- 

1  When  the  coagulins  predominate  they  rapidly  condense  the  antigens, 
giving  the  body  time  to  attack  them  bit  by  bit  without  liberating  enough 
poison  at  a  given  moment  to  cause  toxic  or,  at  least,  fatal  symptoms.  The 
lysins  on  the  contrary,  when  they  predominate,  make  their  appearance  as 
the  agents  of  an  inevitable  and  often  fulminating  intoxication,  for  the  body 
has  only  limited  protection  against  the  true  endotoxins  and  the  true  toxins, 
no  more  than  it  has  against  alkaloids,  for  example  "  (Nicolle). 


ANAPHYLAXIS  249 

sensitiveness  is  due  to  a  lysin  which  sets  free  from  the 
bacteria  a  true  endotoxin. 

Infection  and  intoxication  arouse  in  the  body  a  many- 
sided  conflict  between  these  coagulins  and  lysins,  which  are  in 
general  the  good  and  evil  antibodies.  A  lytic  action  may, 
however,  be  salutary  when  it  occurs  slowly,  whereas  it  is  lethal 
when  it  takes  place  abruptly:  under  these  formulae  may  be 
arranged  all  the  facts  mentioned  in  the  course  of  this  chapter. 
"  Although  diametrically  opposed  to  each  other,  as  inevitable 
results  from  their  definition,  immunity  and  supersensitiveness 
may  co-exist  in  the  same  individual,  as  well  as  succeed  each 
other,  often  again  and  again." 

Nicolle's  theory  is  frankly  inclined  to  the  physical  theory  of 
immunity,  without  overlooking  the  intimate  relations  which 
exist  between  the  physical  properties  of  bodies,  and  their 
chemical  constitution.  It  also  sees  in  immunity  phenomena  of 
nutrition;  for  the  body  digests  the  antigens,  and  the  theory 
supposes  simply  that  every  digestive  act  is  due  to  the  successive 
application  of  a  coagulin  and  a  lysin. 


CHAPTER  XIII 

APPLICATIONS  OF  BACTERIOLOGY 

DIAGNOSTIC  METHODS 

Direct  diagnostic  methods — Direct  diagnosis  of  the  microbe — Cultures 
from  the  blood— Examination  of  faeces— Indirect  diagnostic  methods 
— Cytodiagnoses. 

Biological  diagnostic  methods — Agglutination :  specificity  and  group 
agglutinations  :  variations  in  bacteria  from  the  agglutination  point  of 
view — Precipitation :  employment  in  forensic  medicine  and  in  the 
adulteration  of  foods — Applications  to  anthropology  :  confirmation  of 
the  simian  origin  of  man — Complement-fixation  :  first  experiment  of 
Bordet :  clinical  application — Wassermann's  reaction  and  the  sero- 
diagnosis  of  syphilis — The  nature  of  the  substances  coming  into  play 
in  this  reaction — Supcrscnsitive  reactions  :  tuberculin. 

THE  simplest  and  surest  way  to  diagnose  an  infectious 
disease  is  to  demonstrate  the  presence  of  the  specific  microbe, 
/.*.,  direct  bacteriological  diagnosis.  When  this  is  impossible, 
indirect  diagnosis  is  resorted  to,  i.e.,  the  lesions  of  the  tissues 
which  are  constant  accompaniments  of  a  virus  which  is 
invisible  are  sought  for.  The  presence  in  exudates  of  certain 
cell  elements  is  noted,  or  the  body-fluids  and  bacteria  are 
made  to  react  together  specifically  (antibodies  and  antigens) : 
in  the  latter  case  it  is  more  properly  a  case  of  biological 
diagnosis. 

Direct  Diagnostic  Methods. 

The  microbe  is  sought  for  wherever  there  is  a  possibility 
of  finding  it;  blood,  exudates  and  transudates,  pus,  mucous 
discharges,  false  membranes,  ulcers  and  chancres,  sputum, 

250 


APPLICATIONS   OF  BACTERIOLOGY      251 

cerebro-spinal  fluid,  urine,  faeces,  all  may  be  examined.  Such 
observations  are  completed  by  cultivating  and  experimental 
inoculations. 

Direct  Diagnosis  of  the  Microbe. — This  has  to  suffice 
when  it  is  a  microbe  which  cannot  be  cultivated.  It  is 
sufficient  when  the  microbe  has  characteristics  which  cannot  be 
mistaken.  The  sight  of  the  malarial  parasite  of  Laveran, 
of  a  filaria  embryo,  of  a  trypanosome  (in  man)  is  a  certain 
diagnosis.  Medicine  has  profited  by  every  step  in  advance 
in  technique :  the  ultra-microscope  now  permits  us  to  see 
trypanosomes  and  spirochaates  living  and  motile  much  more 
easily  than  with  the  best  microscope. 

Pasteur's  method  of  "seeding"  silk-worms  was  based  on 
direct  diagnosis.  Direct  diagnosis  is  currently  employed  in 
connection  with  sputum  and  false  membranes ;  it  is  completed 
by  culture  and  inoculation.  Inoculation  is  the  rule  when  the 
pneumococcus  is  observed  in  sputum :  subcutaneously  inocu- 
lated it  kills  a  mouse  within  twenty-four  hours :  the  mouse  is 
the  pneumococcus  reagent. 

Blood  Culture. — Blood,  taken  aseptically  from  an  animal 
not  in  a  state  of  active  digestion,  and  kept  free  from  external 
germs,  may  be  kept  indefinitely  without  putrefying.  If 
microbes  develop  in  the  blood  itself  or  in  nutrient  broth 
into  which  it  has  been  put,  it  means  that  these  microbes 
existed  in  the  blood  during  life.  Pasteur's  observations  on 
the  sterility  of  normal  blood  form  the  foundation  of  the 
diagnosis  of  infectious  diseases  by  blood  culture.  The  strepto- 
coccus was  isolated  by  him  for  the  first  time  from  the  blood  of 
women  suffering  from  puerperal  fever. 

It  has  been  proved  by  blood  culture  that  gonorrhoea,  which  is 
usually  quite  a  local  disease,  may  infect  the  blood  with 
gonococci  and  cause  arthritis  and  endocarditis.  In  a  frankly 
acute  pneumonia  it  is  the  rule  to  find  pneumococci  in  the 
blood. 

In  typhoid  fever  blood  cultivations  have  given  results  which 
have  upset  our  conceptions  of  this  disease.  It  used  to  be 
thought  an  exclusively  local  disease  of  the  intestine  and 


252  MICROBES   AND   TOXINS 

the  lymphoid  organs :  even  when  the  bacillus  had  been  found 
in  the  urine  and  in  the  red  eruption  spots  on  the  skin  it  was 
still  thought  that  it  only  exceptionally  entered  the  blood.  But 
when  cultures  were  regularly  made  from  the  blood  of  typhoid 
patients  it  was  perceived  that  the  typhoid  bacillus  is  present 
there  during  the  whole  febrile  period  of  the  disease  and  again 
during  relapses  :  in  typhoid  fever,  therefore,  the  acute  enteritis 
is  complicated  by  a  blood  infection,  a  septicaemia. 

The  examination  of  the  faeces  is  regularly  performed  in  the 
diagnosis  and  prophylaxis  of  cholera,  typhoid  fever,  and 
dysentery,  particularly  in  order  to  detect  germ-carriers.  Also 
in  the  stools  are  sought  the  eggs  of  worm  parasites  and  of 
ankylostoma  and  amoebae.  The  bacteriological  examination 
of  faeces  is  becoming  more  and  more  common  in  proportion 
as  our  knowledge  of  the  intestinal  flora  is  increasing;  the 
composition  of  this  flora  is  a  guide  to  the  state  of  digestion  and 
nutrition,  and  aids  the  physician  in  his  choice  of  a  diet, 
especially  in  infants,  who  are  so  often  threatened  by  infections 
of  the  alimentary  canal. 

Indirect  Diagnostic  Methods. 

Even  when  no  microbe  is  found  in  it,  the  blood  may  furnish 
diagnostic  proofs  from  its  leucocytic  formula. 

Cytodiagnosis  (Widal)  depends  on  the  examination  of  the 
cells  floating  in  pleural  effusions  or  cerebro-spinal  fluids. 
Different  cells  are  found  in  a  pleurisy  due  to  the  bacillus 
tuberculosis  and  in  one  due  to  heart  disease.  Cytodiagnosis 
of  the  cerebro-spinal  fluid  is  an  indirect  diagnostic  method  in 
certain  tuberculous  and  syphilitic  affections. 

Biological  Methods :  Agglutination. 

The  first  clinical  sero-diagnostic  method  was  discovered  by 
Widal. 

When  a  drop  of  a  broth  culture  of  the  typhoid  bacillus  is 
looked  at  under  the  microscope,  the  bacteria  are  seen  actively 
motile,  isolated  from  eaeh  other  and  dispersed  regularly 


APPLICATIONS   OF  BACTERIOLOGY      253 


throughout  the  liquid ;  the  suspension  is  "  homogeneous."  If 
a  trace  of  serum  is  added  from  an  animal  prepared  by  injections 
of  typhoid  bacilli  or  from  a  patient  suffering  from  typhoid 
fever,  the  bacilli  lose  their 
motility  and  collect  into 
masses :  they  are  said  to 
become  agglutinated  by  the 
serum.  If  the  serum  is 
added  to  a  broth  culture 
floccules  can  be  seen  with 
the  naked  eye  forming  and 
sinking  to  the  bottom  of 
the  tube ;  agglutination  is 
also  a  sedimentation.  Nor- 
mal serum  never  possesses 

this      property,      certainly      FlG<  7I._Aggiutination  of  the  typhoid 
never  to  the  same  degree.  bacillus  by  the  serum  of  a  typhoid 

The  agglutinating  power 
may  be  measured  by  try- 
ing the  effect  on  a  suspension  of  the  bacilli  of  various  dilutions 
of  the  serum :  one  may  say  that  such  and  such  a  serum 
agglutinates  at  i  in  50,  i  in  100,  i  in  1000.  .  .  . 

The  reaction  is  capable  of  two  applications.  With  a  bacillus 
definitely  known  as  a  genuine  B.  typhosus,  one  can  say  that 
the  serum  which  agglutinates  it  is  an  antityphoid  serum.  If, 
on  the  other  hand,  with  such  a  definitely  known  serum  we 
find  a  bacillus  agglutinated,  we  may  say  that  it  is  a  true 
B.  typhosus.  Agglutination  may  be  used  to  diagnose  now  the 
bacillus,  now  the  disease.  The  agglutinating  power  depends 
on  an  antibody  named  the  agglutinin.  This  substance  keeps 
much  longer  than  the  time  required  for  its  carriage  to  long 
distances  for  examination,  when  this  is  necessary.  Dead 
bacteria  also  agglutinate,  so  that  the  method  may  be  employed 
even  without  living  cultures.  Seroagglutination  is  therefore 
the  simplest  and  most  convenient  of  the  biological  methods. 

In  performing  the  test  it  is  necessary  to  avoid  certain 
sources  of  error,  among  others  the  existence  of  bacterial  strains 


254  MICROBES   AND  TOXINS 

which  agglutinate  spontaneously  or  do  not  agglutinate  at  all. 
Strains  can  be  artificially  produced  which  refuse  to  agglutinate  : 
this  resistance  of  the  microbe  to  the  action  of  the  serum 
is  an  example  of  the  immunity  of  the  microbe  towards  the 
animal,  the  converse  case  of  the  immunity  of  the  animal 
towards  the  microbe. 

Agglutination  is  applied  to  as  a  test  for  suspected  bacteria 
found  during  cholera  scares  in  suspected  water  or  choleraic 
diarrhoea ;  the  serum  employed  is  obtained  from  an  animal 
prepared  by  several  immunizing  injections  of  a  definitely  known 
cholera  vibrio. 

Agglutination  has  also  been  applied  to  the  diagnosis  and 
prophylaxis  of  bacillary  dysentery  and  epidemic  cerebro-spinal 
meningitis.  In  tuberculosis  it  has  only  been  applied  as  a 
means  of  controlling  the  treatment  by  tuberculin.  As  a 
diagnostic  agent  the  tuberculin  test  is  much  more  convenient 
and  certain  (R.  Koch).  But  agglutinin  is  no  better  than  the 
other  antibodies  (vide  p.  208)  for  the  estimation  of  the  resist- 
ance of  the  body  towards  tubercle.1 

Precipitation. 

If  we  take  a  culture  of  B.  typhosus  and  filter  it,  we  obtain 
a  clear  fluid  free  from  bacteria.  If  a  little  of  a  very  active 

1  Agglutination  is  the  touchstone  in  the  study  of  bacterial  strains  and 
their  variations.  Recently  Bordet  and  Sleeswyk,  working  with  the  bacillus 
of  whooping-cough  discovered  by  Bordet,  have  created  in  the  laboratory 
varieties  of  this  analogous  to  the  varieties  of  the  dysentery  bacillus. 

What  fixity  do  these  varieties  or  strains  possess,  created  as  they  are  by 
growth  on  different  media  and  separated  by  their  different  reaction  to 
such-and-such  a  serum  ?  What  is  of  interest  here  is  the  conclusion  of  these 
workers.  They  maintain,  at  least  as  far  as  the  agglutinating  action  of 
serum  on  bacteria  is  concerned,  that  sera  "  do  not  act  on  the  fundamental 
bacterial  substance,  which  is  inherent  to  its  life  and  whose  presence  is 
indissolubly  bound  with  the  nature  and  constitution  of  the  species,  but 
that  they  act  on  substances  to  some  degree  accessory,  of  possible  but 
facultative  presence,  whose  production  is  in  no  way  one  of  the  bundle  of 
hereditary,  immutable  characters  which  give  to  a  living  creature  its  own 
physiognomy  and  autonomy." 

It  is  obvious  that  this  interpretation  of  the  facts  is  not  in  favour  of  a 
chemical  theory  of  immunity  :  the  more  one  considers  the  agglutination  as 
a  surface  reaction,  the  more  probable  becomes  the  physical  explanation,  i.e., 
the  explanation  of  Bordet. 


APPLICATIONS   OF  BACTERIOLOGY      255 

antityphoid  serum  is  added,  the  mixture  becomes  turbid  and  a 
precipitate  forms  which  settles  at  the  bottom  of  the  tube. 
Instead  of  agglutination  we  have  precipitation  (R.  Kraus). 

If  we  inject  a  rabbit  repeatedly  with  proteins  of  animal 
origin,  horse  serum,  eel's  serum,  defibrinated  fowl's  blood,  its 
serum  thus  immunized  forms  a  precipitate  when  mixed  with 
the  serum  of  the  eel,  of  the  horse,  or  of  the  fowl  (Tchistovitch 
and  Bordet). 

The  precipitating  property  of  sera  thus  prepared  is  ascribed 
to  an  antibody,  predpitin.  There  are  bacterial  precipitins 
and  protein  precipitins,  the  former  being  simply  a  species  of 
the  latter  :  all  proteins,  animal  or  vegetable,  in  clear  solution,  are 
precipitated  by  the  corresponding  antisera  :  the  action  is  quite 
specific. 

This  reaction  is  nearly  related  to  the  agglutinating  reaction. 
It  is  scarcely  ever  used  in  the  diagnosis  of  infections,  but  for 
the  differentiation  of  protein  substances  it  has  furnished  a  very 
sensitive  test  which  is  employed  in  forensic  medicine  :  nowa- 
days serodiagnosis  is  applied  to  the  detection  of  bloodstains 
and  to  the  adulteration  of  meat  and  milk. 

A  man  is  accused  of  murder  and  in  his  house  is  found  a 
garment  stained  with  blood  :  the  accused  (he  may  be,  for 
example,  a  butcher)  declares  that  it  is  ox-blood  and  justice 
demands  that  serodiagnosis  be  applied  to  the  stains.  Some  of 
the  stains  are  therefore  cut  out  of  the  cloth  and  extracted  with 
physiological  saline  and  with  this  liquid  the  precipitation  test 
is  applied  :  if  the  spot  was  due  to  human  blood,  the  liquid 
produces  a  precipitate  with  the  serum  of  an  animal  previously 
treated  with  human  blood. 

A  merchant  is  accused  of  selling  for  pork-sausages  sausages 
made  from  horseflesh.  An  extract  is  therefore  made  with 
water  from  the  suspected  meat.  If  the  sausage  contained 
horseflesh,  the  extract  gives  a  precipitate  with  the  serum  of  an 
animal  previously  injected  with  horseflesh  extract.  The 
method  can  still  be  applied  when  the  meat  has  been  smoked 
and  dried.  By  the  same  method  may  be  detected  cow's, 
goat's  milk,  &c.  The  reaction  is  sensitive  to  i  in  100,000,  i>., 


256  MICROBES    AND  TOXINS 

it  gives  a  result  with  an  extract  containing  only  y^^ny  of 
its  weight  of  the  protein  to  be  determined. 

The  precipitation  test  has  given  positive  results  with  the 
organs  of  forty-year-old  mummies.  It  is  even  said  to  have 
been  successful  with  mummies  of  3,000  to  5,000  years,  but 
in  this  case  there  is  some  doubt.  It  is  probable  that  these 
latter  may  contain  albumoses,  but  probably  they  no  longer 
contain  coagulable  albumins. 

The  same  biological  reaction  has  furnished  an  additional 
proof  of  evolution.  Just  as  there  are  group  agglutinations,  so 
there  are  group  precipitations.  The  antiserum  which  pre- 
cipitates rabbit  serum  precipitates  also  hare  serum.  Anti-dog 
serum  precipitates  with  both  dog  and  fox  serum.  The  serum 
against  the  horse  precipitates  also  the  serum  of  the  ass  and  of 
the  tapir.  The  relationship,  the  blood  relationship  literally, 
between  the  goat  and  the  sheep,  between  the  fowl  and  the 
pigeon,  and  between  the  domestic  and  wild  pig  can  thus  be 
demonstrated. 

What  interests  us  more  particularly  is  that  the  serum  of  a 
rabbit  prepared  with  human  serum  precipitates  with  the  serum 
of  monkeys,  a  definite  proof  that  we  are  their  near  relations. 
This  relationship  extends  even  to  the  lemurs,  which  are  only 
half-monkeys,  and  is  closer  in  the  case  of  the  monkeys  of  the 
Old  World  than  with  those  of  America.  Griinbaum,  who 
experimented  with  forty-six  species  of  monkeys,  maintains  that 
from  the  point  of  view  of  the  quality  and  quantity  of  the 
precipitates  furnished  by  their  sera,  he  has  been  able  to  detect 
no  difference  between  man,  the  gorilla,  the  orang-outang,  and 
the  chimpanzee :  this  new  proof  of  the  correctness  of  man's 
simian  origin  may  be  welcomed. 

Complement  Fixation. 

This  new  sero-diagnostic  took  origin  from  an  experiment  of 
Bordet :  he  was  trying  to  show  that  there  is  in  a  serum, 
not  several,  but  a  single  complement  and  that  it  is  the  same 
complement  which  acts  in  bacteriolysis  as  in  haemolysis. 


APPLICATIONS   OF   BACTERIOLOGY      257 

We  know  that  complement  (*.*.,  fresh  normal  serum)  is  fixed 
by  the  combination  antigen-antibody.  Let  us  prepare  two 
such  combinations : 

(1)  Cholera  vibrios  and  anticholera  serum  (heated  to  56*  C. 
to  destroy  its  complement). 

(2)  Blood     corpuscles     and     haemolytic     immune     body 
(haemolytic  immune  serum  heated  to  56°  C.). 

Now  on  the  first  combination  fix  complement  (*>.,  add  fresh 
serum  to  the  mixture  and  put  in  the  incubator  :  the  complement 
exerts  its  bacteriolytic  action). 

Now  add  the  second  combination  ready  for  haemolysis  and 
return  to  the  incubator.  If  the  serum  contains  haemolytic 
complement  over  and  above  the  bacteriolytic,  its  presence 
will  be  shown  by  the  laking  of  the  corpuscles  and  the 
red  tint  spreading  throughout  the  mixture.  If,  on  the  other 
hand,  no  haemolysis  occurs,  it  means  that  the  first  fixation 
has  used  up  all  the  complement  of  the  fresh  serum  :  there 
is  no  haemolytic  complement  therefore  distinct  from  the 
bacteriolytic. 

Given  a  mixture  complement  +  immune-body  (antibody) 
+  antigen,  we  have  always  a  means  of  knowing  if  the  com- 
plement has  been  fixed,  namely,  to  add  a  mixture  of  blood 
corpuscles  and  haemolytic  immune-body  ready  for  haemolysis. 
Laking  of  the  blood  indicates  the  absence  of  a  previous 
fixation  of  the  complement.  For  example,  a  serum  is  sent  to 
us  of  a  man  who  has  been  suffering  for  some  time  from  fever ; 
typhoid  fever  is  suspected  ;  if  this  is  the  case  his  blood  ought 
to  contain  an  antibody  (antityphoid  immune-body)  and  in  the 
mixture  of  complement  +  serum  of  the  patient  +  typhoid 
bacilli,  the  complement  will  be  fixed  on  the  bacilli  through  the 
intermediation  of  the  serum.  If  after  a  suitable  lapse  of  time 
one  adds  to  the  same  tube  a  mixture  of  haemolytic  antibody 
and  red  corpuscles,  the  complement  being  no  longer  free,  the 
corpuscles  are  not  dissolved. 

Bordet's  reaction  furnishes  therefore  a  means  of  recognising 
by  the  presence  or  absence  of  haemolysis  the  presence  or 
absence  of  another  immune-body  (antibody),  and  in  conse- 


258  MICROBES  AND  TOXINS 

quence  of  discovering  the  effect  remaining  in  the  animal  body 
from  a  previous  infection. 

It  has  been  used  like  agglutination  to  diagnose  both  a  past 
infection  and  the  nature  of  a  bacterial  strain.  There  is  no 
reason  to  use  it  in  the  diagnosis  of  typhoid  fever,  Widal's 
reaction  being  much  more  convenient.  It  is  very  useful  in 
those  cases  where  agglutination  cannot  be  employed. 

The  Sero-diagnosis  of  Syphilis:  Wassermann 
Reaction. — Wassermann's  reaction  is  simply  the  application 
of  the  Bordet-Gengou  reaction  to  discover  the  syphilitic 
antibody  in  the  serum  of  patients  affected  by  this  malady. 
In  practice  it  is  rather  delicate,  and  demands  an  experienced 
worker.  In  theory  nowadays  it  is  regarded  as  a  precipitating 
reaction  between  certain  colloid  substances  of  the  serum  to  be 
examined  and  the  colloids  of  an  organ  extract  which  is 
employed  as  "  antigen." 

The  reaction  is  not  strictly  specific ;  in  leprosy,  in  trypano- 
soma  infections  in  animals,  perhaps  in  scarlatina,  modifications 
of  the  serum  often  occur  which  give  a  positive  reaction.  The 
method  is  nevertheless  capable  of  clinical  application,  since 
the  physician  has  only  to  decide  between  a  trypanosoma 
infection,  such  as  sleeping  sickness,  which  scarcely  exists  in 
Europe,  leprosy,  which  produces  characteristic  lesions,  scarlet 
fever,  which  is  easy  to  diagnose,  and  syphilis,  which  is 
extremely  common;  the  selection  is  therefore  easy.  A 
laboratory  reaction  which,  taken  by  itself,  does  not  give  an 
absolute  diagnosis,  becomes  nevertheless  an  absolutely  certain 
sign  when  taken  along  with  the  sum-total  of  clinical  and 
biological  indications. 

As  a  matter  of  fact,  the  Wassermann  test,  if  carried  out  with 
the  best  technique,  is  positive  in  about  90  per  cent,  of  the 
cases  of  primary  syphilis,  in  100  per  cent,  in  secondary  syphilis, 
and  in  50-60  per  cent,  of  late  tertiary  cases  (Citron's  figures) ; 
the  proportions  are  rather  lower  in  the  statistics  hitherto  collected 
by  the  Pasteur  Institute.  The  reaction  becomes  more  definite 
the  more  acute  the  infection  and  the  more  active  the  virus. 

There  is  some  doubt  as  to  whether  it  indicates  syphilis  in 


APPLICATIONS   OF  BACTERIOLOGY      259 

full  activity  or  rather  only  that  the  individual  has  been  affected 
at  a  more  or  less  distant  period,  and  that  the  infection  is  now 
latent.  According  to  certain  observers,  it  is  always  the  sign  of 
active  infection,  although  in  a  part  of  the  body  where  the 
lesions  are  not  perceptible,  the  proof  being  that  specific  treat- 
ment makes  the  reaction  disappear,  whereas  it  reappears 
during  relapses  or  recurrences  of  the  disease.  The  experience 
hitherto  with  "  606  "  does  not  apparently  authorize  such  a 
simple  interpretation,  however. 

In  many  cases  the  diagnosis  of  syphilis  is  only  too  easy  j  the 
lesions  visible  and  active  are  sufficient  proof.  But  humanity 
suffers  from  many  ills  which  are  more  or  less  distant  con- 
sequences of  an  attack  of  syphilis,  neglected,  forgotten,  or 
even  unsuspected.  Since  syphilis  is  a  curable  disease  the  value  of 
Wassermann's  reaction  in  the  diagnosis  of  these  obscure  cases 
is  obvious  :  lesions  of  the  liver,  of  the  heart,  of  the  aorta,  and 
failure  of  the  sight  may  all  by  its  means  be  assigned  to  their 
true  cause  and  yield  to  the  specific  treatment.  An  old  clinical 
law  holds  that  the  mother  of  a  syphilitic  infant,  without  being 
herself  infected,  is  immune  to  infection  from  her  child,  and 
thus  may  nurse  it  without  danger ;  in  reality,  it  turns  out,  she  is 
infected  :  the  serum  reaction  proves  it.  A  Dresden  physician, 
Rietschel,  has  thus  diagnosed  syphilis  in  10  per  cent,  of  wet 
nurses  apparently  healthy,  and  the  confirmation  was  obtained 
when  it  was  found  that  the  children  of  these  women  eventually 
showed  signs  of  syphilis. 

Syphilis  unsuspected  or  without  symptoms  is  a  disease  less 
rare  than  is  generally  thought,  and  Wassermann's  reaction 
furnishes  us  with  an  additional  weapon  against  this  formidable 
enemy. 

Clinical  medicine  has  long  maintained  the  syphilitic  origin 
of  general  paralysis  of  the  insane,  and  of  locomotor  ataxy. 
The  sero-diagnosis  brings  a  striking  proof  of  this.  Not  only 
the  serum  but  the  cerebro-spinal  fluid  of  general  paralytics 
furnishes  a  positive  reaction  in  88  per  cent,  of  all  cases,  and  in 
95  per  cent,  of  advanced  cases.  In  locomotor  ataxy  the  per- 
centage is  not  so  high. 

S  2 


260  MICROBES   AND   TOXINS 

The  reaction  is  also  of  value  in  the  distinction  between 
general  paralysis  and  other  mental  affections  which  bear  an 
external  resemblance  to  it,  among  others  from  dementia  praecox. 
Bordet's  reaction  has  also  been  applied  in  the  diagnosis  of 
hydatid  cysts  due  to  the  echinococcus. 

Also  it  has  been  employed  in  the  detection  of  blood  and 
foreign  proteins  for  which  it  is  an  even  more  sensitive  test  than 
precipitation  :  it  requires,  however,  such  delicate  manipulation, 
and  is  so  extremely  sensitive,  that  it  can  hardly  be  trusted  in 
certain  cases;  according  to  Friedberger  it  is  capable  of 
detecting  the  minute  trace  of  protein  present  in  human  per- 
spiration, so  that  the  hands  of  the  experimenter  himself  may 
introduce  an  error. 

It  has  been  employed  in  the  differentiation  of  different  races 
of  man  and  of  monkeys  :  not  that  the  distinction  between  man 
and  monkeys  is  so  absolutely  necessary  as  to  show  the  sort  of 
hierarchy  which  exists.  Man  is  as  far  removed  from  the  orang 
as  the  orang  is  from  the  macacus  ;  however,  man  is  more  nearly 
related  to  the  orang  than  the  latter  is  to  certain  species  of 
macacus.  It  is  even  possible,  it  is  said,  to  distinguish  by  sero- 
diagnosis  an  individual  of  Mongolian  race  from  a  Malay. 

The  biological  reactions  thus  are  of  value,  not  only  in  the 
physiology  of  immunity  and  of  nutrition,  but  even  in  zoology 
and  anthropology. 

Tuberculin  Tests. 

A  dose  of  tuberculin  which  produces  no  effect  in  a  healthy 
subject  may  determine  in  a  tuberculous  individual  a  definite 
reaction,  consisting  of  general  and  local  phenomena,  fever, 
inflammation,  and  oedema  round  the  tubercles  and  at  the  site 
of  injection. 

It  was  long  thought  that  the  reaction  only  took  place  in  the 
neighbourhood  of  the  tuberculous  lesions  themselves,  and 
consisted  in  the  inflammation  and  destruction  of  a  certain 
number  of  cells,  the  fever  being  due  to  the  absorption  of  the 
cellular  debris.  Von  Pirquet,  however,  discovered  that  the 
skin  of  a  tuberculous  patient  reacts  to  tuberculin  at  every  point, 


APPLICATIONS   OF  BACTERIOLOGY      261 

and  it  has  therefore  been  necessary  to  suppose  that  the  body  is 
impregnated  throughout  by  some  substance  produced  by  the 
chronic  infection. 

The  body  responds  to  the  attack  of  the  tubercle  bacillus  by 
a  reaction  product,  an  antibody,  and  the  tuberculin  reaction 
appears  to  be  produced  by  the  coming  together  or  combination 
(not  necessarily  in  the  chemical  sense)  of  this  antibody  and 
tuberculin ;  that  is  roughly  how  the  facts  are  being  interpreted 
nowadays. 

The  body  by  producing  this  antibody  is  said  to  become 
"  sensitive  "  to  tuberculin.  The  tuberculous  individual  behaves 
differently  from  the  healthy  subject,  being  more  sensitive,  and 
his  excessive  susceptibility  acting  as  a  warning  to  him  of  his 
danger.  It  is  in  this  way  that  the  supersensitiveness  which  is 
a  phase  of  immunity  (v.  Chap.  XII.,  page  240)  has  occasionally 
been  interpreted.  There  is  some  resemblance  between  the 
tuberculin  reaction  and  the  anaphylactic  shock  ;  the  diagnosis 
of  tuberculosis  by  tuberculin  is  based  on  the  "  supersensitive- 
ness "  of  the  individual. 

The  reaction  represents  in  a  sense  a  resistance  to  the  action 
of  tuberculin ;  we  have  seen  that  the  prototype  of  this  reaction, 
Koch's  phenomenon,  consists  in  the  expulsion  of  the  re- 
inoculated  virus  by  the  infected  subject ;  it  is  therefore  also  an 
immunity  reaction. 

Tuberculin  was  at  first  employed  as  a  remedy,  but  later  only 
as  a  diagnostic  agent.  It  was  even  more  studied  in  veterinary 
science  than  in  medicine,  because  it  was  easier  to  make  experi- 
ments and  control  them  by  autopsies  on  animals.  Even  to-day 
the  tuberculin  test  is  quite  as  important  in  veterinary  as  in 
human  practice. 

In  the  diagnosis  of  tubercles,  nowadays,  smaller  doses  are 
used  for  subcutaneous  injections  than  at  the  time  of  the  first 
trials  of  tuberculin. 

The  temperature  reaches  its  greatest  height  about  eight 
hours  after  injection.  A  positive  reaction  indicates  merely 
"tuberculous  lesions"  and  nothing  more.  It  gives  no 
information  as  to  the  site  or  the  age  of  the  lesions,  nor 


262  MICROBES  AND  TOXINS 

as  to  the  probable  fate  of  the  patient.  The  reaction  may  be 
negative  in  exhausted,  cachectic  tuberculous  patients. 

The  number  of  individuals  who  give  a  positive  reaction 
much  exceeds  the  number  of  those  who  show  clinical  signs  of 
tubercle :  latent  dormant  tuberculosis,  or  even  healing  tubercle, 
gives  a  reaction  because  the  body  is  saturated  with  the 
products  of  its  reaction  to  the  tuberculous  infection.1 

In  veterinary  practice,  it  happens  that  animals  once  sub- 
jected to  tuberculin  injection  fail  to  react  to  a  second  test 
performed  twenty-five  to  thirty  days  after  the  first ;  this  fact 
has  been  employed  in  fraudulent  dealing  to  throw  out  the 
diagnosis,  but  the  second  test  will  still  act  if  a  double  dose 
is  given,  and  if  the  temperature  is  taken  at  the  fourth 
hour  (instead  of  the  tenth);  the  fever  appears  prematurely 
(Vallee). 

Cutaneous  Reactions. — In  individuals  affected  by  tuber- 
culosis, if  the  skin  is  subjected  to  a  quite  superficial  scarifica- 
tion on  which  is  put  a  drop  of  tuberculin,  a  red  spot  is  seen  to 
appear  after  a  few  hours.  This  swells,  and  comes  to  resemble 
a  little  vaccine  pustule.  It  was  this  phenomenon  which  was 
discovered  by  Von  Pirquet.  Several  variations  have  been 
tried.  It  is  possible  to  do  without  scarification  by  dropping 
into  the  inner  corner  of  the  eye  a  drop  of  a  dilute  solution  of 
tuberculin;  the  conjunctiva  reacts  with  more  or  less  intense 
inflammation  and  more  or  less  exudate ;  this  is  the  ophthalmo- 
reaction  of  Wolff-Eisner  and  of  Calmette. 

It  was  observed  during  subcutaneous  injections  of  tuberculin, 
that  the  needle-track  of  the  syringe,  being  wetted  with  a  trace 
of  the  tuberculin,  became  red  and  swelled  up ;  from  this 

1  The  statistics  of  Franz  dealing  with  certain  regiments  of  the  Austro- 
Hungarian  army  are  worth  reproducing,  since  the  author  had  the 
opportunity  of  observing  the  same  men  during  several  years. 

In  a  regiment  of  four  hundred  apparently  healthy  men  recruited  from  a 
district  where  tuberculosis  was  rife,  61  per  cent,  gave  during  their  first  year 
of  service  a  positive  reaction  ;  of  323  similar  men  in  their  second  year  of 
service  68  per  cent,  were  positive.  Of  279  healthy  men  recruited  from  a 
district  where  tuberculosis  was  scarce,  there  were  30  per  cent,  of  positive 
reaction.  These  1002  individuals  furnished  in  the  course  of  the  seven 
following  years  sixty-four  cases  of  manifest  tubercle  and  twenty  died  of  it. 


APPLICATIONS   OF  BACTERIOLOGY      263 

subordinate  phenomenon  an  independent  diagnostic  test  has 
been  established  by  making  a  simple  prick  in  the  skin  (Moro). 

Further,  it  is  possible  to  inject  into  the  thickness  of  the 
skin  itself  a  drop  of  the  solution  containing  only  yj^  of  a 
milligram  of  tuberculin ;  there  is  thus  obtained  a  very  definite 
diagnostic  reaction  :  the  intradermo-reaction  (Mantoux). 

These  procedures  are  all  very  convenient,  especially  in  the 
examination  of  children ;  the  methods  of  pricking  and  of 
intradermo-reaction  are  less  severe  than  the  intra-ocular 
instillation. 

The  cutaneous  reaction  of  V.  Pirquet  gives  about  85-90  per 
cent,  of  positive  results  in  children  presenting  signs  of  tuber- 
culosis, 20  per  cent,  in  children  with  no  clinical  symptoms,  and 
48  per  cent,  in  doubtful  cases.  The  percentage  of  positive 
reactions  in  children  not  clinically  tuberculous  varies  with  the 
age.  During  the  first  6  months  of  life  it  is  practically  o ; 
from  6  to  24  months  2  per  cent. ;  from  2  to  4  years  13  per 
cent.  ;  from  4  to  six  years  17  per  cent.  ;  from  6  to  10  years 
35  per  cent.  •  from  10  to  14  years  55  per  cent.  ;  in  adults 
there  are  at  least  77  per  cent,  of  positive  reactions. 

The  great  value  of  V.  Pirquet's  cutaneous  reaction  is  that 
with  a  little  prick,  which  is  itself  quite  harmless  and  produces 
no  trace  of  fever,  and  with  a  minimal  quantity  of  tuberculin 
(down  to  Y^Vir  °f  a  milligram),  it  is  possible  to  tell  if  a  child  is 
affected  with  tubercle  bacillus. 

The  younger  the  child  the  more  definitely  does  the 
reaction  signify  a  true  active  infection,  since  tuberculosis  is  less 
frequent — and  more  recent — the  younger  the  infant.  In  the 
adult,  it  indicates  for  the  most  part  a  tuberculous  impregnation, 
great  or  small,  recent  or  remote. 

The  cutaneous  tuberculin  reaction,  performed  as  it  has  been 
on  thousands  and  thousands  of  human  beings,  has  taught  us 
that  man  becomes  infected  with  tubercle  in  childhood  from  his 
first  to  his  fourteenth  year;  so  that  tuberculosis,  although  it 
may  only  become  serious  and  fatal  at  a  more  advanced  age,  is 
really  a  disease  of  children  and  not  of  adult  life.  It  is  then 
during  childhood  that  protection  is  necessary  and  since  we 


264  MICROBES   AND  TOXINS 

know  that  the  child  is  infected  by  the  individuals  surrounding 
it,  it  is  in  the  family  itself  and  in  the  home  that  contagion 
must  be  prevented.  The  disease  is  kept  up  and  spread  by 
infected  members  of  the  family  and  by  servants  and  also  by 
unhealthy  houses,  in  particular  by  dark,  airless  rooms.  The 
cutaneous  reaction  tells  us  that  of  a  hundred  adults  harbouring 
the  tubercle  bacillus,  only  twelve  are  destined  to  die  of 
tubercle.  It  is  therefore  very  evident  that  many  human 
beings  recover  from  the  attack  of  the  bacillus  and  that  man  is 
an  animal  naturally  resistant  to  tuberculosis  ;  in  most  of  us  a 
sort  of  spontaneous  vaccination  takes  place.  These  facts 
ought  not  to  be  overlooked  in  the  search  for  a  method  of 
treatment  or  prevention  of  tubercle. 


CHAPTER  XIV 

VACCINES   AND   SERA 

Vaccination  with  unknown  viruses — Small-pox  :  inoculation  and  Jennerian 
vaccination — Sheep-pox — Rabies  :  Pasteur's  treatment — Vaccination 
with  microbes  of  attenuated  virulence — Anthrax,  Swine-erysipelas, 
Pleuropneumonia  of  cattle— Vaccines  against  cholera,  plague,  typhoid 
fever — Attempts  at  anti-tuberculous  vaccination. 

Bacteriotheraphy :  intestinal,  buccal,  nasal — Applications  to  surgery. 
Serotherapy,  Diphtheria,  Tetanus,  Venoms,  Cholera — Serum  against 
plague,  bacillary  dysentery,  cerebrospinal  meningitis —  Anti-strepto- 
coccic  sera — Anti-tuberculous  sera. 

Sero-vaccinations  :  Sheep-pox — Bovine  plague — Swine-plague. 

Virus-vaccines  sensitized— Experiments  of  Ehrlich  and  Morgenroth. 
Besredka's  vaccines :  Typhoid  fever,  Plague — Virus-serum  in  the 
treatment  of  rabies. 

Phagocytic  therapy. 

IT  is  fairly  easy  to  immunise  laboratory  animals  against 
bacteria  by  repeated  doses  of  the  living  microbe :  a  preliminary 
experiment  determines  the  minimum  lethal  dose,  and  the  first 
injections  are  to  be  kept  well  below  this.  Immunization  is 
possible  also  with  viruses  which  are  incapable  of  killing  in 
any  dose. 

With  extremely  virulent  bacteria,  immunization  is  much 
more  difficult.  It  is  impossible  to  immunize  a  guinea-pig 
against  anthrax,  for  example.  In  man,  where  one  is  not 
entitled  to  run  the  slightest  risk,  it  is  impossible  to  begin  even 
with  very  small  doses  of  living  virulent  microbes.  Hence 
arises  the  great  importance  of  Pasteur's  discovery  in  the  history 
of  active  immunization,  the  discovery  of  the  attenuation  of  # 
virus.  Only  vaccines  may  be  used  in  human  immunization. 

In   general,  a  non-virulent  microbe   neither  kills   nor  im- 

265 


266  MICROBES   AND  TOXINS 

munises,  whereas  a  virulent  organism  kills  before  immunity 
has  time  to  develop.  It  is  often  necessary  to  find  inter- 
mediate stages  of  virulence.  This  is  what  Pasteur  perceived, 
and  hence  arose  his  two  vaccines  against  anthrax. 

It  is  possible  to  vaccinate  with  viruses  which  one  handles 
without  actually  knowing  their  nature,  with  cultures  of  bacteria 
virulent  or  attenuated,  and  with  bacterial  products  freed  from 
the  living  microbes. 

VACCINES 
I.   Vaccinations  with  Unknown  Viruses. 

It  is  not  necessary  actually  to  know  the  microbe  of  a 
disease  in  order  to  immunize  against  it.  The  vaccinations 
with  unknown  viruses  are,  as  a  matter  of  fact,  among  the  most 
perfect  known  to  medicine. 

Small-pox. — The  microbe  of  small-pox  is  unknown,  yet 
preventive  inoculations  have  long  been  carried  on.  There 
exist  two  methods,  both  empirical  in  origin ;  the  older  of  the 
two  has  been  entirely  given  up  in  favour  of  Jenner's  method 
of  vaccination.  Inoculation  or  variolisation  consisted  in 
inoculating  the  virus  of  small-pox  itself,  so  as  to  produce  one 
or  more  cutaneous  pustules,  the  development  of  which  saved 
the  body  from  a  general  attack.  Small-pox  acquired  in  the 
natural  manner  invades  the  whole  body :  the  virus  may  enter 
perhaps  by  the  tonsils  or  the  respiratory  passages,  gaining  the 
blood  stream  from  these  and  thence  reaching  the  skin  and 
mucous  membranes.  Once  the  eruption  has  appeared  it  is 
too  late  to  intervene.  When  the  natural  virus  is  inoculated 
artificially  on  the  skin,  it  may  give  rise  to  a  general  infection 
later  on,  but  in  general  the  first  local  pustules  produce 
rapidly  sufficient  immunity  to  prevent  such  a  general  invasion ; 
but  not  always;  occasionally  inoculation  results  in  a  fatal 
attack  of  small-pox.  The  virus  inoculated  was  by  no  means 
constant,  and  it  was  always  a  case  of  working  in  the  dark. 
A  correct  and  witty  account  of  the  history  of  inoculation  is  to 
be  found  in  the  eleventh  of  Voltaire's  Lettres  Philosophiques. 


VACCINES   AND   SERA  267 

There  exists  among  cattle  a  disease  resembling  small-pox, 
and  also  due  to  an  unknown  microbe,  the  cow-pox^  which 
attacks  also  those  who  milk  infected  cows.  Those  individuals 
who  have  suffered  from  the  pustules  of  cow-pox  are  immune  to 
small-pox. 

This  idea,  popular  in  origin,  had  long  been  current  among 
farmers  and  cattle-breeders  in  certain  districts  of  England, 
France,  Holland,  and  Germany  when  Jenner  took  it  up 
and  consecrated  his  life  and  his  genius  to  publishing  it 
abroad. 

He  made  of  the  subject  a  definite  experimental  study :  he 
inoculated  in  turn  the  virus  from  animal  to  animal,  from  the 
animal  to  man,  from  man  to  man,  and  finally  submitted  his 
"  vaccinated  "  subjects  to  experimental  inoculation  with  small- 
pox. 

In  all  countries  where  public  health  is  well  organized  vacci- 
nation has  been  made  compulsory  ;  in  consequence  smallpox 
has  almost  disappeared  from  Germany  :  the  rare  cases  recorded 
there  are  cases  of  immigrants  or  inhabitants  on  the  frontiers. 
It  has  very  much  diminished  in  France  and  is  being  actively 
combated  in  the  French  colonies.  It  may  one  day  become 
extinct. 

Three  improvements  have  been  introduced  in  Jennerian 
vaccination  since  Jenner's  time.  In  the  first  place,  it  was 
recognized  that  in  many  cases  vaccination  does  not  protect  for 
the  whole  of  life,  and  periodical  revaccinations  have  become 
customary  and  even  in  certain  countries  compulsory.  In  the 
second  place,  instead  of  vaccinating  from  man  to  man  the 
vaccine  employed  is  derived  from  animals.  Jenner  practised 
"  arm  to  arm "  vaccination,  i.e.,  transmitted  from  man  to 
man  a  virus  whose  distant  origin  was  cowpox.  This  practice 
had  the  inconvenience  of  occasionally  transmitting  human 
diseases  such  as  syphilis.  From  1860  onwards  vaccination  in 
Europe  has  been  performed  with  "  animal  lymph "  taken 
from  vaccine  calves  (in  certain  colonies  from  buffalos,  rabbits, 
zebras,  &c.). 

Finally  we  have  learned  to  preserve  and  purify  the  supply  of 


268  MICROBES   AND   TOXINS 

virus  by  mixing  it  with  glycerine,  so  that  it  is  no  longer  necessary 
to  vaccinate  directly  from  the  calf.  Inflamed  arms  are  nowa- 
days quite  the  exception. 

According  to  certain  observers  there  are  really  only  two  types 
of  smallpox,  the  pox  of  sheep  and  that  of  man.  All  the  others 
are  said  to  be  mere  modifications  of  these  two  types ;  for  small- 
pox is  very  apt  to  become  modified  in  adapting  itself  to  different 
animal  species. 

Vaccinia,  in  this  theory,  would  simply  be  an  adaptation  of 
human  smallpox  to  cattle,  and,  starting  with  this  idea,  attempts 
have  been  made  to  transform  smallpox  into  cowpox  experiment- 
ally by  inoculating  from  man  to  the  calf.  This  question  of 
variolo-vaccinia  has  been  very  much  studied  and  is  being  taken 
up  again  to-day.  The  results  obtained  have  been  much  dis- 
cussed, and  it  is  not  yet  absolutely  proved  that  it  is  possible  to 
produce  in  the  laboratory  in  a  short  time  a  change  which 
nature  has  probably  required  centuries  to  accomplish. 

Rabies. — Pasteur's  treatment  takes  advantage  of  the  period 
of  incubation,  which  is  on  the  average  six  weeks.  The  patient 
ought  to  be  treated  as  soon  as  possible  after  the  bite  and  before 
the  appearance  of  symptoms. 

In  the  process  there  is  inoculated  the  virus  fixe,  which  is 
kept  going  in  the  nervous  system  of  the  rabbit,  is  contained  in 
the  spinal  cord,  and  may  be  modified  by  drying  the  latter  in 
the  dark  at  the  temperature  of  23°.  To  begin  with,  a  cord  dried 
for  fourteen  days  is  injected,  and  at  the  end  of  the  treatment  a 
cord  dried  for  three  days.  The  rate  of  treatment  varies  accord- 
ing to  the  site  and  gravity  of  the  bites,  there  being  three 
formulae,  the  fifteen-day,  eighteen-day,  and  twenty-one  day 
treatments.  The  mortality  in  rabies  has  been  reduced  to  0-3 
per  cent,  instead  of  15  to  20  per  cent. 

The  microbe  of  rabies  is  unknown,  and  Pasteur  treatment 
is  one  of  the  crowning  glories  of  experimental  medicine. 
Consider  the  facts  which  it  has  been  necessary  to  determine 
before  arriving  at  its  application  :  in  the  first  place,  the  virus 
of  rabies  exists  in  the  brain  of  rabid  animals  ;  further,  it  is 
possible  to  make  a  pure  inoculation  of  it ;  the  virus  of  any 


VACCINES   AND   SERA  269 

rabid  dog  taken  at  random  inoculated  under  the  dura  mater 
of  the  brain  of  a  rabbit  after  trephining  the  skull  produces 
rabies  in  the  rabbit  after  a  variable  period  of  incubation ;  but 
after  about  twenty  passages  from  rabbit  to  rabbit  the  incuba- 
tion period  falls  to  six,  to  seven  days  ;  the  virus  has  then 
become  the  virus  fixe ',  and  the  passages  from  rabbit  to  rabbit 
are  really  cultures  in  the  living  body.  The  culture  exposed  to 
dry  air,  but  sheltered  from  light,  gradually  loses  some  of  its 
virulence.  The  dried  cords,  inoculated  in  the  form  of  emulsions 
under  the  skin  of  animals,  produce  in  them  a  certain  and 
stable  immunity  against  the  strongest  virus  inoculated  under 
the  dura  mater.  The  cultivation  in  vivo  of  the  virus  fxe  was 
for  Pasteur  the  key  to  vaccination  against  rabies  after  the  bite 
of  the  rabid  animal. 

There  is  some  doubt  whether  the  virus  of  the  dried  cords 
really  undergoes  attenuation.  Even  in  1885  Pasteur  said  that  it 
was  rather  a  rarefaction  which  occurred.  "  The  delays  observed 
in  the  duration  of  incubation  of  the  rabies  communicated  day 
by  day  to  rabbits,  to  estimate  the  virulence  of  our  cprds  dried 
in  contact  with  air,  result  from  a  diminution  in  quantity  of  the 
virus  contained,  and  not  from  a  diminution  in  virulence."  A 
variation  of  Pasteur's  treatment,  the  treatment  of  Hogyes, 
consists  in  replacing  the  graduated  desiccation  by  a  graduated 
dilution.1 

1  The  nature  of  this  virus  fixe  cultivated  in  the  rabbit  is  rather  a 
question.  Nitsch  and  Marx,  who  inoculated  themselves  with  fresh  virus 
fixe,  consider  that  after  several  hundred  passages  in  rabbits  the  virus 
becomes  harmless  to  man  ;  they  consider  it  as  a  true  Jennerian  vaccine  for 
man,  resulting  by  adaptation  to  a  different  species.  But  it  is  not  right  to 
maintain  the  absolute  innocuousness  of  -virus  fixe.  It  may  well  be  harm- 
less when  injected  into  the  subcutaneous  tissue,  but  would  it  continue  to  be 
so  if  injected  in  a  region  rich  in  nerve  filaments  such  as  the  face  or  the  end 
of  finger  ? 

It  is  probable  that  the  virus  of  dried  cords  is  destroyed  by  the  body,  that 
the  microbe  is  absorbed,  and  that  protective  substances  are  produced  in 
consequence  and  are  found  in  the  blood  of  treated  individuals.  But  these 
substances  play  only  a  secondary  part,  for  there  is  no  constant  relation 
between  the  properties  of  the  serum  of  an  animal  and  its  resistance  to 
rabies.  There  are  certain  animals  refractory  to  rabies,  such  as  a  species  of 
tortoise  whose  blood  nevertheless  possesses  no  antirabic  power.  It  is 
impossible  to  avoid  thinking  of  the  phagocytes  in  such  a  condition. 


270  MICROBES   AND  TOXINS 


ii.    Vaccination  with  Attenuated  Cultures. 

Anthrax. — The  anthrax  vaccines  are  cultures  attenuated 
by  contact  with  the  air  at  a  temperature  of  42*5°  C.  Two 
graduated  vaccines  are  employed :  the  weaker  kills  mice  and 
small  guinea-pigs,  the  stronger  kills  adult  guinea-pigs  and  even 
a  certain  percentage  of  rabbits.  The  animals  treated  with 
these  preserve  their  immunity  for  about  a  year,  and  it  has  been 
calculated  that  during  the  twenty  years  which  followed  their 
introduction  at  least  twelve  million  animals  were  inoculated. 
The  method  of  anthrax  vaccination  has  never  required  alter- 
ation since  its  discovery  by  Pasteur,  Chamberland,  and  Roux, 
and  the  celebrated  experiment  of  Pouilly-le-Fort. 

Swine-Erysipelas. — In  this  the  same  principle  is 
employed,  /.<?.,  a  virus  attenuated  in  the  laboratory.  But  the 
bacillus  of  swine-erysipelas  does  not  produce  spores,  and  a 
method  of  sero-vaccination  has  been  invented  instead  (v.  infra). 


Hi.  Pleuropneumonia  of  Cattle  (Rinderpest). 

The  discovery  of  the  microbe  dates  only  from  1898,  but 
vaccination  had  been  practised  against  the  disease  a  con- 
siderable time  before.  Willems  employed  as  virus  the  serous 
fluid  from  the  diseased  lungs  :  he  had  observed  that  inoculation 
of  this  in  a  healthy  animal  produced  variable  effects  according 
to  the  site  of  injection  chosen  :  injected  on  the  trunk  or  neck 
the  infection  was  fatal ;  at  the  tip  of  the  tail  or  of  the  ears  it 
produced  only  a  limited  inflammation  which  left  the  animal 
immune  towards  the  naturally  occurring  disease. 

The  first  improvement  was  introduced  by  Pasteur,  who 
showed  that  the  pure  virus  could  be  got  by  taking  the  abundant 
exudate  produced  in  the  subcutaneous  tissue  of  a  calf  after 
inoculation  there.  Later,  pure  cultures  were  substituted. 

The  tip  of  the  tail  is  a  good  site  for  vaccination,  since  the 
tissue  is  dense  and  the  absorption  slow.  A  few  animals  lose 
their  tails  as  a  result  of  the  vaccination. 


VACCINES  AND  SERA  271 


iv.  Cholera,  Typhoid  Fever,  Plague. 

Cholera. — Human  cholera  is  a  toxic  disease,  the  toxin 
being  secreted  by  the  vibrios  invading  the  intestine.  The  true 
remedy  for  cholera,  therefore,  should  be  an  antitoxic  serum. 
It  was  before  our  present  conceptions  as  to  the  toxicity  of  the 
cholera  vibrio  that  Ferran,  in  1884,  tried  to  vaccinate  men 
against  cholera  by  injecting  the  living  microbe  subcutaneously. 
This  method  has  been  taken  up  and  modified  by  Haffkine, 
who  has  tried  it  in  India,  where  cholera  is  always  present. 
In  vaccinated  individuals  he  found  the  mortality  seven  times 
less  than  in  the  non-vaccinated,  but  although  the  death-rate 
may  be  lower,  the  severity  of  the  symptoms  is  quite  as  great 
among  the  former  as  among  the  latter,  which  is  rather  dis- 
concerting, as  in  other  kinds  of  preventive  vaccination  this 
does  not  occur.  The  solution  of  the  cholera  problem  lies 
elsewhere,  but  it  is  worth  while  recalling  the  attempts  of  Ferran 
and  Haffkine,  because  their  method  has  proved  of  more  value 
in  other  diseases. 

Typhoid  Fever. — Typhoid  fever  is  a  toxic  enteritis,  but 
it  is  at  the  same  time  a  blood-infection  ;  blood-cultures  show 
that  there  is  septicaemia.  The  bacillus  inhabits  chiefly  the 
small  intestine,  but  it  also  extends  to  the  blood,  the  spleen, 
and  the  bone-marrow.  Hence  there  is  much  more  hope  of 
success  than  with  cholera  for  preventive  injections  of  the 
bacteria  under  the  skin.  Bacilli  killed  by  heat  are  employed. 
The  injection  produces  some  swelling  and  pain  with  fever  and 
stiffness,  the  discomfort  lasting  about  two  days.  Two,  or  even 
three,  injections  ought  to  be  made.  The  treatment  is  succeeded 
by  what  is  called  a  "negative  phase,"  during  which  the 
individual  may  be  in  a  condition  of  lowered  resistance. 

Anti-typhoid  vaccination  has  been  chiefly  employed  hitherto 
in  the  English  army  in  India,  in  the  Transvaal,  in  Egypt,  and 
in  Cyprus.  A  certain  number  of  the  inoculated  individuals 
have  nevertheless  taken  the  disease  (in  many  of  these  the 
vaccination  had  been  insufficient),  but  in  a  form  milder  and 


MICROBES   AND  TOXINS 

with  less  frequent  relapses.  The  mortality  is  about  four  times 
less  among  the  inoculated  than  among  the  non-inoculated. 
As  far  as  can  be  judged  at  present,  the  effect  of  the  vaccination 
lasts  for  about  four  years. 

There  is  no  necessity  to  submit  a  whole  family  to  the  dis- 
comforts of  inoculation  when  one  member  has  taken  typhoid ; 
it  is  sufficient  to  isolate  the  patient  and  to  disinfect  strictly 
the  excreta,  linen,  etc.  Vaccination  is  adapted  only  for  the 
members  of  an  ambulance  and  hospital  corps  during  an 
epidemic,  or  for  troops  campaigning  in  a  country  where 
typhoid  is  raging,  and  where  it  is  difficult  to  adopt  hygienic 
measures. 

Plague. — Haffkine's  vaccine  consists  of  bacteria  killed  by 
heat.  The  inoculations,  especially  the  first,  produce  inflamma- 
tion and  pain  with  a  high  temperature,  the  illness  lasting  for 
about  three  days.  Two,  or  even  three,  inoculations  ought  to 
be  made.  Such  vaccinations  are  suitable  in  the  case  of  plague, 
as  this  disease  is  a  general  infection,  spreading  throughout  the 
blood  and  organs.  In  1902-1903  about  half  a  million  persons 
were  inoculated  in  the  Punjab,  and  fairly  accurate  informa- 
tion could  be  gathered  regarding  a  portion  of  them.  On 
comparison  we  find  : — 

186,797         inoculated,   with  3339    cases  =  I '8  per  cent. 
,,       814  deaths  =  0-4  per  cent. 

639,630  non-inoculated,  with  49,433   cases  =  7*7  per  cent. 
»    29,733  deaths  =  47  per  cent. 

It  appears  that  the  inoculation  continues  to  be  of  benefit  for 
several  years  after. 

Anti-tuberculous  Vaccination.— As  far  as  is  known, 
vaccination  against  tubercle  is  possible  with  living  bacilli  only, 
dead  bacilli  being  ineffective,  but  there  is  no  vaccine  as  yet 
which  is  entirely  safe. 

Behring  has  sought  "  Jennerian  "  vaccines,  i.e.,  he  has  tried 
to  vaccinate  one  species,  cattle,  with  the  virus  derived  from 
another  species,  man.  Innumerable  experiments  have  already 
been  made  on  bovo-vaccination,  and  they  prove  at  least  that 
a  certain  degree  of  immunity  may  be  created  towards  tuber- 


VACCINES   AND   SERA  273 

culosis.     But  even  among  cattle  the  method  is  not  yet  suited 
for  practical  application. 

In  man  there  exists  an  auto-vaccination,  since  practically  all 
adults  have  been  slightly  affected  by  this  disease,  and  only  a 
small  fraction  succumb.  Both  in  medical  and  surgical  practice 
it  is  common  to  find  tuberculosis  of  the  skin,  of  the  joints,  and 
of  the  bones  healing  up  and  even  apparently  protecting  the 
patients  against  contracting  pulmonary  phthisis.  It  is  on  these 
lines  that  is  to  be  sought  a  method  of  vaccination  against 
tuberculosis.  Up  to  now  none  exists. 

BACTERIOTHERAPY. 

Vaccines  and  sera  are  not  the  only  remedies  furnished  to 
medicine  by  bacteriology.  Bacteriotherapy  consists  in  modify- 
ing a  given  flora  by  adding  to  it  other  bacteria  which  are 
antagonistic  or  favourable  to  certain  microbial  species.  In 
discussing  the  intestinal  flora  we  have  indicated  the  principles 
of  the  bacteriotherapy  of  intestinal  infections.  In  the  future 
this  will  be  developed,  and  health-giving  bacteria  will  form 
part  of  our  diets. 

Bacteriotherapy  has  also  been  attempted  in  the  case  of 
cancer  and  lupus,  where  injections  of  streptococci  have  been 
given,  and  in  the  case  of  chronic  boils,  where  injections  of 
yeast  have  been  tried.  These  attempts  are  still  semi-empirical, 
but  this  is  no  reason  for  despising  them. 

The  lactic  bacilli  have  not  been  utilized  in  intestinal  bacterio- 
therapy only.  Surgery  has  employed  them  to  combat  puer- 
peral infections,  injuries  occurring  during  labour,  inflammations 
of  the  cervix,  and  even  peritonitis.  They  evidently  act  by 
modifying  the  "  soil."  It  is  obvious  that  they  find  their  chief 
success  in  combating  infections  of  closed  cavities,  such  as  we 
have  in  rhinitis,  sinusitis,  ethmoiditis,  and  otitis. 

Bacteriotherapy  in  the  mouth  is  not  less  rational  than  in 
the  intestine.  The  lactic  flora  has  succeeded  in  curing 
chronic  inflammations  of  the  gums  and  obstinate  pyorrhoea 
alveolaris.  All  the  infectious  diseases  which  enter  by  the 


274  MICROBES   AND  TOXINS 

mouth,  the  tonsils,  and  the  nasopharynx  may  be  combated  by 
this  bacteriotherapy,  and  already  very  encouraging  observations 
exist.  For  example,  epidemic  cerebro-spinal  meningitis  starts 
from  the  nasopharynx,  and  there  are  meningococcus  carriers 
just  as  there  are  diphtheria  carriers.  It  is  the  more  important 
to  employ  the  lactic  bacilli  against  the  meningococcus  from  the 
fact  that  in  vitro  the  former  antagonize  the  latter  (A.  Berthelot). 
The  bacterium  most  to  be  recommended  for  the  purpose  is 
the  Bacillus  bulgaricus  administered  in  powders  and  derived 
only  from  young  fresh  cultures  in  liquid  media.  It  is  not 
necessary  to  insist  upon  the  inconvenience  of  the  use  of  anti- 
septic washes  and  to  contrast  with  these  the  biological 
disinfection  produced  by  modifying  the  flora. 

SERA. 

Serotherapy  means  treatment  by  means  of  the  sera  of 
immunized  animals.  It  may  be  preventive  or  curative,  or  both. 
The  animal  furnishing  the  serum  has  produced  its  own 
immunity,  t.e.,  it  possesses  active  immunity.  The  animal 
which  receives  such  a  serum  acquires  a  passive  immunity. 
Serotheraphy  is  equivalent  to  the  transfusion  of  antibodies. 

The  antitoxins  were  discovered  by  Behring  in  1890. 

Antitoxic  sera  are  not  the  only  therapeutic  sera:  by 
immunizing  an  animal  with  the  bodies  of  bacteria  antibacterial 
serum  can  be  prepared :  with  such  sera  there  may  be 
transfused  bacteriolysins,  agglutinins,  bacteriotropins,  in  fact 
all  the  antibodies  they  contain,  but  above  all  there  is  transferred 
their  property  of  exciting  the  reaction  of  the  phagocytes. 

Certain  antibacterial  sera  are  also  anti-endotoxic  in  the 
sense  that  they  are  capable  of  neutralizing  endotoxins.  They 
are  best  prepared  as  we  have  seen  by  intravenous  injections  of 
the  antigen.  Finally,  sera  may  be  prepared  against  viruses 
only  slightly  known  or  not  known  at  all ;  the  preventive  action 
of  serum  in  general  may  be  disregarded. 

Antidiphtheria  Serum. — This  is  a  serum  of  horses 
immunised  against  the  diphtheria  toxin.  The  treatment  of  the 


VACCINES  AND   SERA  275 

horse  begins  not  with  pure  toxin  but  with  a  mixture  of  toxin  + 
Gram's  iodine  or  of  toxin  +  antitoxin.  Every  horse  has  its 
own  susceptibility,  and  the  treatment  requires  a  good  deal  of 
skill.  It  is  impossible  to  foretell  the  supply  of  antitoxin  from 
any  particular  horse. 

The  serum  treated  aseptically  is  put  into  little  bottles  and 
"tyndallised,"  i.e.,  sterilised  by  heating  to  a  low  temperature 
on  several  successive  days.  It  is  stored  in  a  dark,  cool  place, 
and  under  these  conditions  it  only  loses  about  one-tenth  of  its 
activity,  so  long  as  it  remains  clear,  even  after  several  years. 

Sera  possessing  great  activity  per  unit  volume  have  been 
sought  for  by  selecting  those  horses  which  are  active  pro- 
ducers, or  by  concentrating  or  "refining"  the  serum  by 
various  procedures,  freezing,  precipitation,  &c.  The  best 
method  is  still  to  take  an  active  serum,  to  keep  it  pure  and 
preserve  it  carefully  without  further  treatment. 

The  titration  of  the  serum  is  indispensable :  only  serum  of 
known  activity  is  issued  for  serum  therapy.  Hitherto,  titration 
has  only  been  possible  by  resorting  to  experimental  animals, 
guinea-pigs.  The  laboratories  of  all  countries  have  agreed 
to  adopt  the  method  proposed  by  Ehrlich.  The  serum  is 
standardized  by  mixing  with  a  toxin  which  has  itself  been 
estimated  against  a  standard  antitoxin.  The  activity  of  the 
serum  is  expressed  in  antitoxic  units  which,  like  all  units  of 
measure,  are  conventional.  The  standard  antitoxin  is  preserved 
with  precautions  similar  to  those  with  which  are  guarded  our 
standard  weights  and  measures. 

But  in  spite  of  all  the  precision  introduced,  we  are  still 
dealing  here  with  a  biological,  not  a  chemical  process. 
Experience  has  made  clear  three  points  on  which  too  much 
stress  cannot  be  laid,  (i)  The  serum  ought  to  be  inoculated 
as  soon  as  possible  after  the  appearance  of  the  disease.  A 
little  serum  given  early  is  worth  more  than  a  great  deal  of 
serum  given  too  late.  Similar  guinea-pigs  receiving  the  same 
dose  of  toxin  may  be  cured  by  a  small  dose  at  the  sixth 
hour,  whereas  at  the  eighth  hour  even  a  large  dose  fails.  In 
hospital,  unfortunately,  the  children  are  usually  brought  too 

T  2 


276  MICROBES   AND  TOXINS 

late,  and  it  is  on  this  account  that  the  mortality  in  diphtheria  is 
still  10  per  cent,  (or  14  per  cent,  in  epidemics ;  L.  Martin). 

(2).  There  should  be  no  fear  of  giving  large  doses :  in  fact 
in  bad  throats  large  doses  ought  to  be  resorted  to  at  once. 

(3).  Intravenous  injection  is  the  most  efficacious  :  it  is 
indicated  in  all  serious  cases.  This  holds  good  for  all  sera. 

There  is  said  to  exist  still  in  certain  minds  a  certain 
scepticism  as  to  the  value  of  antidiphtheritic  serum  but  it  is 
scarcely  credible.  The  figures  speak  f@r  themselves.  The 
death  rate  from  diphtheria  in  the  Sick  Children's  Hospital  in 
Paris  before  serotherapy  was  about  50  per  cent.;  it  has  fallen 
to  6 — 10  per  cent.,  and  would  be  much  less  if  the  children  were 
treated  at  the  very  beginning  of  the  disease. 

The  serum  is  used  as  a  prophylactic  against  diphtheria  in 
schools,  barracks,  creches  and  hospitals :  a  dose  of  5 — 10  c.c. 
protects  for  about  three  weeks. 

Anti-tetanic  Serum  is  an  antitoxic  prophylactic  serum. 
The  serum  furnished  by  the  Pasteur  Institute  possesses  an 
activity  such  that  roirVsu-  c-c-  sun<ices  to  neutralize  in  vitro 
100  lethal  doses  for  a  mouse.1 

In  human  tetanus  when  the  incubation  is  long  and  the 
progress  slow,  the  serum  may  check  the  extension,  i.e.,  may 
have  a  limiting  effect.  According  to  the  figures  collected  by 
Vallas  the  serum  has  lowered  the  death  rate  from  about  75 
per  cent,  to  45  per  cent.  The  mortality  remains  about  70 
per  cent,  in  those  cases  where  the  incubation  period  is  less 
than  eight  days. 

1  When  tetanus  toxin  and  a  sufficient  dose  of  antitoxin  are  injected  at 
different  points  of  the  body  a  minute  quantity  of  toxin  escapes  neutraliza- 
tion and  induces  a  slight  local  tetanus  which  is  always  recovered  from. 

It  is  possible  to  cure  animals  inoculated  in  the  paw  if  the  serum  is 
injected  at  the  first  appearance  of  symptoms  :  when  the  poison  has  reached 
the  nerve-centres  the  antitoxin  is  of  no  avail.  Roux  and  Borrel  had  the 
idea  of  bringing  the  antitoxin  in  direct  contact  with  the  brain  so  as  to 
prevent  the  poison  reaching  the  centres.  By  intracerebral  inoculation 
they  succeeded  in  curing  a  number  of  guinea-pigs  which  had  manifested 
tetanus  and  were  therefore  doomed  to  die.  But  they  do  not  extend  their 
conclusions  to  other  animals.  In  man  the  first  symptoms  of  tetanus 
affect  always  the  medullary  centres  so  that  cerebral  treatment  has  even  less 
hope  of  success  than  in  the  guinea-pig. 


VACCINES   AND   SERA  277 

The  employment  of  this  serum  in  veterinary  practice  is  due 
to  the  efforts  of  Nocard.  He  collected,  now  a  long  time  ago, 
2,708  cases  of  horses  injected  immediately  after  one  of  those 
operations  frequently  followed  by  tetanus  (castration,  amputa- 
tion of  the  tail,  inguinal  or  umbilical  hernia) ;  not  a  single  case 
of  tetanus  occurred.  A  second  group  comprising  600  animals, 
treated  one  to  four  or  even  more  days  after  an  accidental 
wound  (nail-prick,  harrow-tooth  wound,  kicks,  wounds  soiled 
with  earth  or  manure,  etc.),  presented  one  single  case  of  mild 
tetanus  (a  horse  treated  five  days  after  the  accident,  a  nail- 
prick  in  shoeing).  The  veterinary  surgeons  who  supplied  these 
3,308  cases  observed  in  their  practice  at  the  same  time  314 
cases  of  tetanus  (of  which  220  were  horses)  in  animals  not 
treated  with  serum.  In  705  equidae,  wounded  or  operated 
on,  Labat  observed  three  cases  of  tetanus  among  the  un- 
treated, not  a  single  case  among  the  treated  animals.  Accord- 
ing to  statistics  supplied  by  eight  veterinary  surgeons  to 
Vallee  of  Alfort,  from  1898  to  1906,  13,126  animals  were 
inoculated  as  a  preventive  measure,  and  not  one  took  tetanus. 
During  the  same  period,  two  of  these  surgeons  alone  had 
139  cases  of  tetanus  among  animals  not  treated.  At  present 
the  Pasteur  Institute  supplies  to  veterinary  surgeons  more  than 
100,000  doses  per  annum. 

There  is  no  reason  known  to  science  why  man  should  not 
derive  a  similar  advantage.  There  exists,  however,  with 
regard  to  the  prophylactic  serotherapy  of  tetanus  in  man  a 
distrust  or  scepticism  which  finds  vent  from  time  to  time  in 
the  discussions  of  surgical  societies  or  congresses.  It  has 
been  maintained  that  the  employment  of  preventive  injections 
has  not  affected  the  death  rate  from  tetanus  in  Paris  ;  that 
tetanus  occurs  fairly  often  in  spite  of  prophylactic  injections ; 
that  the  conditions  which  permit  of  the  prophylactic  employ- 
ment in  veterinary  practice  are  unrealizable  in  human  medicine, 
and  finally  that  in  man  the  serum  is  less  active  than  in  the 
horse.  When  examined  none  of  these  objections  hold  good. 
The  numerous  unsuccessful  cases  depend  on  the  conditions  of 
tetanus  intoxication  and  its  neutralization  -9  there  is  not  one  of 


278  MICROBES   AND  TOXINS 

i 

the  objectors  who,  if  he  received  a  tetanus-suspected  wound, 
would  not  call  for  a  prophylactic  injection  of  serum  for 
himself. 

It  is  necessary  to  avoid  giving  too  small  doses  and,  should 
the  wound  fail  to  heal,  to  repeat  the  injection  several  times ; 
of  course,  the  surgical  cleansing  of  the  wound  should  never  be 
omitted. 

An ti venom  Sera. — These  are  prepared  by  immunizing 
horses,  beginning  with  venom  +  calcium  hypochlorite  and  ending 
with  pure  venom.  Eventually  they  can  be  made  to  support 
80  lethal  doses  (2  grams  of  dried  venom)  at  a  single  injection. 
The  immunization  is  a  difficult  operation  to  bring  to  a  satis- 
factory end ;  frequently  it  is  interrupted  by  serious  accidents, 
endocarditis,  nephritis,  and  abscesses.  It  takes  16  months 
of  treatment  to  get  a  good  serum  from  a  horse  (Calmette). 
Anti venom  serum  is  polyvalent,  i.e.,  is  prepared  with  several 
species  of  venom. 

The  quantity  of  serum  required  to  save  life  increases  with 
the  susceptibility  of  the  animal  and  the  delay  in  the  application 
of  the  remedy.  As  a  rule  10-20  c.c.  suffice  to  save  a  bitten 
human  being. 

Anticholera  Serum. — Cholera  is  an  acute  intoxication 
due  to  a  poison  elaborated  by  the  vibrios  in  the  intestine. 
This  toxin  has  been  sought  for  and  an  antitoxic  serum 
attempted.  The  serum  prepared  against  the  bacteria  is  not 
active  against  the  poison ;  the  principle  of  serotherapy  is 
thus  quite  different  from  the  Ferran-Haffkine  method  of 
immunization. 

The  cholera  toxin  prepared  in  1896  by  Metchnikoff,  Roux, 
and  Salimbeni  presented  this  peculiarity,  that  it  resisted  heating 
to  100°  C.  for  a  quarter  of  an  hour.  Horses  are  prepared  by 
intravenous  injections. 

Anticholera  serum  is  still  at  the  trial  stage.  Its  only  trial  out- 
side the  laboratory  was  in  1909,  during  the  epidemic  of  St. 
Petersburg.  The  doses  ought  to  be  large,  150-350  c.c.,  given 
100  c.c.  at  a  time  intravenously.  There  are  cases  so  fulminant 
as  to  defy  almost  any  remedy.  In  the  above  trial  cases,  not 


VACCINES   AND   SERA  279 

very  numerous,  the  mortality  was  reduced  by  about  a  half.  In 
the  laboratories  work  is  still  going  on  towards  the  improvement 
and  perfection  of  an  anticholera  serum. 

Antiplague  Serum. — It  is  derived  from  horses  immunized 
against  the  plague  bacillus.  The  treatment  is  long  and  difficult. 

The  statistics  are  in  need  of  discussion.  Whereas  Mayer, 
for  example,  records  numerous  cases  of  cure,  other  physicians 
declare  that  there  are  more  deaths  among  the  treated  than  the 
non-treated :  they  allege  that  the  serum  is  too  weak  and  the 
Hindu  too  susceptible  to  plague.  When  these  unfavourable 
statistics  are  more  closely  examined,  it  becomes  apparent  that 
the  serum  has  been  given  too  late,  in  too  small  dose,  by  subcu- 
taneous injection,  and  not  long  enough  continued :  the  patients, 
abandoned  after  the  fall  of  the  fever,  died  in  many  cases 
fifteen  to  thirty  days  after  the  last  serum  injection.  In  man, 
plague  is  all  over  in  five  to  six  days,  and  if  treatment  is  only 
carried  on  during  the  third  or  fourth  day  cure  is  well  nigh 
impossible. 

The  injections  ought  to  be  massive  and  repeated.  It  is  no 
use  trying  to  treat  plague  with  the  doses  suitable  to  antitetanus 
serum.1 

It  is  evident  that  against  pneumonic  plague  the  serum  which 
we  possess  at  present  is  not  to  be  relied  on,  but  that  against 
bubonic  plague  it  has  already  been  of  service.  It  may  be  of 
great  value  in  a  population  where  the  hygiene  is  good,  to 
complete  the  isolation  of  a  case  of  plague  by  giving  prophylactic 
injections  to  the  contact  individuals,  and  thus  to  limit  the 
epidemic  focus. 

Antidysenteric  serum  is  given  in  doses  of  20,  80  to 
100  c.c. — the  latter  doses  in  very  serious  cases  where  there  are 
more  than  100  motions  of  the  bowel  per  day.  Between  1905 

1  Duprat,  using  doses  of  300  c.c.,  succeeded  in  curing  thirty-eight  out  of 
forty -five  plague  patients  treated,  a  mortality  of  only  15  per  cent. 

Employing  intravenous  injections  (100  c.c.  repeated  after  twenty-four 
hours),  Penna  had  a  mortality  of  14*2  per  cent,  in  200  cases.  Ferrari  at 
Sao  Sabastiao  had  only  7*2  per  cent,  of  fatal  cases  among  forty-four. 
Ferrari's  statistics  throw  light  on  the  different  modes  of  injection :  by  the 
.skin,  38  per  cent,  death-rate,  intraperitoneally,  18  per  cent,  intravenously 
7*2  per  cent. 


280  MICROBES   AND  TOXINS 

and  1907  Vaillard  and  Dopter  applied  the  serum  in  512  acute 
cases  of  all  ages,  of  whom  32  had  certainly  fatal  dysentery 
and  170  a  very  severe  attack.  The  average  death  rate  was 
only  i -3  per  cent.,  whereas  in  different  epidemics  it  has  varied 
from  10  to  25  or  50  per  cent.  The  serum  relieves  the  patients 
of  their  terrible  abdominal  pains,  of  the  vomiting,  and  of  the 
hiccough ;  the  stools  become  less  numerous  and  less  painful, 
the  temperature  falls,  and  a  feeling  of  comfort  succeeds  that 
of  absolute  prostration.  These  good  results  occur  as  a  rule  in 
the  course  of  one  day. 

The  antimeningococcus  serum  of  Flexner  is  employed 
in  epidemic  cerebro-spinal  meningitis.  It  has  hardly  any 
action  when  injected  subcutaneously.  It  must  be  injected  at 
the  site  of  the  disease — i.e.,  into  the  cerebro-spinal  canal  by 
lumbar  puncture.  The  treatment  should  be  begun  as  soon  as 
possible  and  be  repeated.  It  has  lowered  the  mortality  from 
60  to  80  per  cent,  to  25  to  10  per  cent. 

Antistreptococcic  serum  is  a  polyvalent  serum— i.e.,  it 
is  prepared  with  several  strains  of  streptococci.  It  is  difficult 
to  get  an  accurate  idea  of  its  efficacy  in  man.  The  infections 
in  which  streptococci  are  responsible  are  multiple,  and  the 
serum  is  employed  in  the  most  diverse  diseases,  from  puerperal 
fever  to  scarlatina  and  acute  rheumatism.  Large  quantities  of 
this  serum  are  supplied  from  the  Pasteur  Institute  both  for 
medical  and  veterinary  use,  but  unfortunately  it  is  rare  to  get 
any  information  as  to  the  results  obtained. 

Antituberculous  Sera. — Maragliano's  serum  is  supplied 
by  horses  treated  with  the  soluble  constituents  of  the  tubercle 
bacillus. 

Marmorek's  serum  is  furnished  by  horses  immunized 
with  "primitive  bacilli,"  *>.,  young  bacilli  which  scarcely 
possess  any  of  the  acid-fast  property  due  to  the  wax  and  fat 
constituents.  The  same  horses  are  at  the  same  time  im- 
munized against  streptococci,  since  the  streptococci  are 
frequently  associated  with  Koch's  bacillus  in  tuberculous 
lesions.  As  regards  this  serum  opinions  differ.  Certain 
observers  report  a  favourable  action  on  cases  of  tuberculosis  of 


VACCINES   AND   SERA  281 

the  bones  and  joints  and  on  early  cases  of  pulmonary  tubercle 
(checking  the  fever  and  improving  the  general  condition). 

SERO  VACCINATIONS. 

When  a  vaccination  per  se  involves  a  certain  risk  it  may  be 
associated  with  a  specific  serum :  the  combined  treatment  is 
equivalent  to  an  active  immunization  under  cover  of  a  passive 
protection. 

Sheep-pox.1 — In  France  the  law  enjoins  the  vaccination 
against  sheep-pox  of  all  the  flocks  in  the  affected  regions, 
whereas  it  forbids  it  in  the  regions  free  from  the  scourge.  In 
Algeria  sheep-pox  is  endemic,  and  the  Algerian  sheep  are 
much  more  resistant  than  the  French  sheep.  In  consequence, 
Algerian  sheep,  which  are  imported  in  great  numbers,  may  be 
landed  apparently  healthy,  yet  may  induce  a  malignant 
epidemic  in  a  French  flock.  Nowadays  only  vaccinated  sheep 
may  be  imported. 

Borrel,  who  perfected  the  method  of  vaccination  by  showing 
how  to  prepare  cheaply  large  quantities  of  pure  virus,  has 
prepared  an  active  anti-serum  by  "  charging  "  with  virus  sheep 
which  had  recovered  from  the  disease.  When  injected  into  a 
sheep  twenty-four  hours  before  the  virus  this  serum  prevents 
the  development  of  the  disease  (in  a  dose  of  20-30  c.c.),  and 
even  inhibits  the  inoculation  pustule.  By  graduating  the  dose 
of  serum  it  is  possible  by  inoculating  the  virus  simultaneously 
to  produce  simply  an  immunizing  pustule  without  any  risk  of 
a  general  infection. 

The  serum  employed  alone  is  sufficiently  active  to  render 
certain  a  prophylactic  treatment  of  the  disease  by  curative  and 
preventive  injections  in  a  flock.  It  permits  of  the  importation 
of  Algerian  sheep  without  danger  of  infecting  French  territory. 
The  sheep  to  be  imported  receive  the  serum  three  weeks 
before  shipment.  Inspection  eliminates  those  which  were 
incubating  the  disease  at  the  time  of  inoculation,  and  which 
might  therefore  be  shipped  while  still  ill.  This  serotherapy  is 

1  Non-existent  in  Great  Britain. — Translators'  note. 


282  MICROBES   AND  TOXINS 

of  advantage  to  the  Algerian  breeders,  and  helps  in  the 
suppression  of  the  disease. 

Cattle  plague  or  rinderpest  is  an  epizootic  disease  which 
has  caused  incalculable  losses  throughout  Europe,  especially 
by  attacking  the  herds  of  pure  blood.  The  mortality  always 
exceeds  50  per  cent.  The  microbe  is  unknown,  but  it  is 
known  that  the  virus  can  pass  through  filter  bougies,  even 
the  finest :  it  is  therefore  an  ultramicroscopic  organism.  An 
animal  which  survives  possesses  an  immunity  which  seems 
quite  invincible :  as  soon  as  the  illness  is  over  any  quantity 
of  virulent  blood  may  be  injected  into  it  without  killing  it. 
But  it  is  not  possible  to  immunize  by  a  method  similar  to 
that  in  sheep-pox,  for  inoculation  produces  an  attack  as  severe 
as  the  naturally  acquired  disease. 

The  animals  may  be  made  to  acquire  an  immunity  lasting 
for  four  to  six  months  by  injecting  the  bile  of  animals  dead  of 
the  plague,  but  the  method  is  not  practical. 

A  good  serum  can  be  obtained  by  "  charging  "  with  virus  an 
animal  which  has  recovered.  The  sero-vaccination  of  Kolle 
and  Turner  consists  in  injecting  simultaneously  this  serum  and 
virulent  blood  :  they  must  be  injected  at  different  spots. 

It  is  always  risky  to  employ  a  virus  in  therapeutics  :  there  is 
always  the  danger  of  spreading  it  or  introducing  it  into  a 
country  where  it  does  not  hitherto  exist.  The  serum  alone 
suffices  for  prophylaxis,  for  treatment,  and  for  the  prophylaxis 
of  the  healthy  animals.  The  prophylactic  dose  is  one  of 
50  c.c.  Capable  veterinary  surgeons  have  succeeded  by  using 
the  serum  alone  in  reducing  the  mortality  to  12  per  cent. 
(Nicolle).  The  same  principles  are  at  work  in  the  treatment  of 
the  horse-sickness  of  the  Transvaal  and  the  plague  of  pigs,  hog- 
cholera,  which  are  also  septicaemic  diseases  due  to  an  ultra- 
microscopic  virus. 

SENSITIZED  VACCINES. 

The  vaccines  employed  against  typhoid  fever,  cholera  and 
plague  may  be  rendered  less  painful,  more  prompt,  more 
powerful  and  more  durable  by  impregnating  the  bacteria  of 


VACCINES   AND  SERA  283 

which  they  consist  with  the  corresponding  anti-sera 
(Besredka).  After  soaking  in  the  serum  the  bacteria  are 
washed  free  from  all  trace  of  it :  they  must  only  retain  what 
they  have  been  able  to  fix  :  there  must  be  no  free  serum. 

These  qualities  of  such  vaccines  are  due  to  the  immune 
body  which  they  have  fixed.1 

PHAGOCYTIC  THERAPY. 

When  the  leucocytes  are  counted  to  determine  the 
diagnosis  and  prognosis  in  certain  infectious  diseases  it  is 
really  a  case  of  measuring  to  some  extent  the  reactions  and 
natural  means  of  defence  of  the  body :  the  laws  of  phagocy- 
tosis are  being  applied.  It  was  natural  not  to  stop  at 
recording  these  phenomena,  but  to  proceed  to  active  inter- 
vention by  attempting  to  modify,  fortify  or  direct  the 
phagocytic  apparatus. 

The  great  danger  in  surgical  operations,  especially  on  the 
abdomen,  is  an  infection  of  the  field  of  operation  :  formerly 
it  was  the  custom  to  flood  this  with  antiseptics ;  the  bacteria 
were  destroyed  but  the  tissues  were  injured.  Nowadays  not 
only  has  antisepsis  given  place  to  pure  asepsis  but  the  attempt 
is  made  to  summon  to  the  field  of  operation,  particularly  to 
the  peritoneum,  legions  of  the  cells  capable  of  taking  up 
bacteria  and  healing  wounds.  For  this  purpose  it  is 
customary  to  inject  into  the  abdominal  cavity  either  blood 
serum  (warmed  to  body  temperature),  or  substances  which 

1  The  association  of  an  antirabic  serum  with  the  injection  of  the 
emulsions  of  spinal  cord  may  be  regarded  as  a  sero- vaccination  or  as 
treatment  with  a  sensitized  vaccine. 

Such  a  serum  cannot  be  relied  upon  to  prevent  rabies  in  animals  :  still 
less  can  it  furnish  the  basis  of  treatment  for  man.  But  it  has  been 
employed  with  success  in  conjunction  with  the  virus  fixe  of  the  Pasteur 
treatment  to  induce  the  greatest  possible  absorption  by  the  body.  In  such 
virus-serum  mixtures  it  is  only  the  virus  which  immunizes :  the  serum 
merely  favours  absorption.  An  excess  of  serum  would  be  actually 
injurious  and  must  be  avoided. 

In  man  the  virus-serum  treatment  is  employed  in  those  cases  where  the 
treatment  has  to  be  rapid,  i.e..,  when  the  patient  presents  himself  long  after 
the  bite  or  where  the  injuries  have  been  very  serious  (A.  Marie). 


284  MICROBES  AND  TOXINS 

exert  a  positive  chemiotaxis  on  the  leucocytes,  e.g.,  nucleic 
acid. 

Again,  in  the  method  of  Bier,  which  consists  in  cupping  and 
surrounding  with  elastic  bands  to  retain  a  large  volume  of 
blood  around  an  abscess,  a  boil,  or  other  acute  infection,  it  is 
also  the  phagocytes  which  are  called  upon,  for  the  artificial 
oedema  which  is  produced  is  accompanied  by  an  afflux  of 
leucocytes  (Schimodaira's  experiment). 

Wright's  method  of  vaccino-therapy  consists  in  treating 
infections  by  means  of  inoculations  of  bacterial  bodies  (the 
bacteria  of  the  infection  itself),  in  order  to  increase  the 
opsonic  power  of  the  serum  (v.  Chap.  X.) ;  the  treatment 
would  not  be  applicable  without  the  quantitative  and  quali- 
tative control  in  which  the  phagocytes  are  employed. 

For  some  years  the  study  has  been  going  on  of  those 
chemical  actions  capable  of  stimulating  phagocytosis  and 
reinforcing  the  phagocytes.  Weak  solutions  of  quinine 
(0*002  per  cent.)  increase  the  phagocytic  power,  whereas 
solutions  fifty  times  stronger  produce  the  opposite  effect 
The  peptones  have  been  recognized  to  be  powerful  stimulants. 
According  to  quite  recent  experiments,  it  will  be  necessary  to 
add  to  these  iodoform  and  various  chemical  substances  which 
are  soluble  in  fats,  and  which  doubtless  act  in  virtue  of  their 
consequent  power  of  penetrating  protoplasm  in  solution  in  the 
lipoids.  These  substances  are  true  chemical  stimulins. 

It  is  possible  that  a  new  and  most  interesting  chapter  is 
about  to  open  in  therapeutics  with  the  discovery  of  Hamburger 
among  others :  the  action  of  certain  mineral  springs  is  said 
to  be  due  to  their  power  of  exciting  phagocytosis  (experiment 
with  the  water  of  the  Virchow  spring  in  Wiesbaden). 

Thus,  chemistry  and  physics  are  far  from  taking  possession 
of  medicine  and  relegating  phagocytosis  to  the  region  of  pure 
speculation.  On  the  contrary,  the  doctrine  of  phagocytosis, 
which  has  already  bloomed  a  second  time  in  the  purely 
biological  researches  on  opsonins  and  bacteriotropins,  plays 
the  premier  role  still  as  inspiration  and  explanation  in  the 
study  of  the  chemical  therapeutic  agents. 


CHAPTER  XV 

CHEMICAL  REMEDIES 

CHEMIOTHERAPY 

Ancient  origin  of  chemical  therapeutic  agents — "Sterilization"  of   the 

infected  animal— Protozoal  diseases  :  example  of  quinine — Association 

of  chemical  research   with  animal  experiment. 
The  arsenical  bodies  and  "  606  "  or  Salvarsan. 
Progress  in  the  treatment  of  sleeping-sickness  and  syphilis — Observations 

on  immunity  in  Protozoal  diseases. 
Strains  resistant  to  drugs — Strains  resistant  to  sera — The  future  of  chemio- 

therapy. 

CHEMICAL  therapeutics  is  old  as  the  world.  It  fills  our 
pharmacopoeias,  still  further  enriched  by  organic  chemistry. 
But  all  over  it  furnishes  only  symptomatic  remedies :  one 
induces  sleep,  another  stimulates  the  heart,  others  deaden  pain. 
The  drugs  which  really  cure  are  easily  counted,  few  exist 
besides  quinine  and  mercury. 

Chemical  therapeutics  of  to-day  is  engaged  in  searching 
for  other  examples  of  the  order  of  quinine  and  mercury,  drugs 
capable  of  destroying  pathogenic  microbes  without  injuring  the 
cells  of  the  body.  In  theory  and  practice  it  is  chemistry 
applied  to  therapeutics. 

Such  remedies  should  resemble  more  or  less  the  antiseptics 
and  should  have  as  function  sterilization.  When  the  antiseptic 
properties  of  corrosive  sublimate  became  known,  R.  Koch 
tried  to  inject  it  into  anthrax-infected  guinea-pigs  to  destroy 
the  bacteria,  but  the  animals  died.  This  simple  experiment 
puts  the  problem  well  before  us  with  all  its  difficulties. 
Corrosive  sublimate  cannot  be  employed  to  "  sterilize "  a 

285 


286  MICROBES  AND  TOXINS 

living  creature  because  it  kills  the  cells  of  the  organs  at  the 
same  time  as  the  bacterial  cells.  Quinine  is  a  good  thera- 
peutic agent  because  it  destroys  the  parasites  of  the  red 
corpuscles  without  acting — at  least  in  general  — on  the 
corpuscles  themselves. 

There  are  cases  where  the  sterilization  of  a  tissue  may  be 
performed  without  the  least  inconvenience.  Mercury  is  the 
traditional  remedy  in  syphilis*  its  internal  application  is 
limited  by  its  toxicity  ;  but  after  a  contact  possibly  dangerous, 
it  may  be  applied  as  a  prophylactic  to  the  skin.  Metchnikoffs 
experiments  on  the  chimpanzee  and  on  man  have  proved 
that  simple  rubbing  with  calomel  ointment  sterilises  the  skin 
where  the  microbe  of  syphilis  has  just  been  inoculated  and 
prevents  infection.  This  preventive  treatment,  which  is  as 
efficacious  as  it  is  simple,  is  already  practised. 

Origins. — The  renaissance  of  chemiotherapy  has  had 
several  causes.  The  first  is  the  revolution  due  to  Pasteur's 
teaching.  When  a  disease  is  found  to  be  due  to  a  germ,  to  a 
germ  which  is  known  and  which  can  be  made  to  transmit  the 
disease  by  inoculation  into  a  laboratory  animal,  then  there  is 
room  for  experimental  therapeutics.  Animal  experiment 
permits  us  to  go  much  further  than  clinical  observation. 
Nevertheless,  the  progress  of  microbiology  at  first  seemed  about 
to  throw  drug  therapy  into  the  background.  Marvellous 
biological  remedies  were  discovered,  both  preventive  and 
curative,  surpassing  in  specificity  and  efficacy  all  the  drugs  of 
the  Pharmacopoeia,  e.g.  the  Pasteur  vaccine  against  anthrax, 
vaccination  against  rabies,  and  all  the  serotherapies.  The  idea 
at  once  suggested  itself  that  all  infectious  diseases  might  be 
treated  in  similar  ways. 

But  that  has  not  proved  to  be  the  case ;  there  is  a  group  of 
infectious  diseases  against  which  vaccination  and  serotherapy 
have  achieved  nothing  or  almost  nothing,  the  protozoal  diseases. 
In  no  case  has  the  serum  of  an  animal  susceptible  to  the 
trypanosome  of  sleeping  sickness,  and  immunized  against  this 
microbe,  been  capable  of  exerting  a  curative  effect  upon  the 
experimental  disease  in  other  animals.  In  malaria,  all  the 


CHEMICAL  REMEDIES  287 

biological  methods  have  failed,  and  quinine  remains  the 
sovereign  remedy.  Hence,  instead  of  searching  for  a  vaccine 
or  a  serum  in  protozoal  diseases,  the  search  has  been  directed 
to  finding  an  equivalent  of  quinine  for  each. 

When  fortune  favours,  the  history  of  cinchona  bark  and 
quinine  may  be  recapitulated.  Chance  or  an  empirical  dis- 
covery or  a  tradition  of  unknown  origin,  then  enriches 
humanity  with  a  remedy  on  which  chemists  and  experimenters 
exercise  their  talents.  This  is  the  history  of  the  recent  arsenical 
remedies. 

The  discovery  of  the  microbes  and  the  necessity  of  dis- 
covering for  protozoal  diseases  other  remedies  than  vaccines 
and  serum,  have  been  aided  by  the  development  of  chemistry 
itself,  and  especially  of  industrial  chemistry.  The  great  aniline- 
dye  factories  have  been  the  chief  furnishers  of  the  laboratories 
of  experimental  medicine.  The  remedies  tried  have  been 
varied  as  the  dyes  are  varied,  and  in  many  cases  chemiotherapy 
has  been  really  a  chromotherapy. 

The  method  of  research  consists  in  associating  experiments 
on  the  living  animal  with  the  reactions  of  organic  chemistry. 
A  certain  body  endowed  with  a  certain  property  is  known  ;  the 
molecule  of  this  body  possesses  a  principal  nucleus  on  which 
are  grafted  secondary  atomic  groups;  it  is  found  that  the 
property  desired  depends  upon  one  of  these  groups ;  this  group 
is  then  shifted  about,  varied  or  substituted  by  another  group ; 
experiment  then  informs  us  as  to  the  properties  of  the 
new  compound  and  as  to  the  relations  between  molecular 
structure  and  therapeutic  effect. 

Ehrlich  has  rendered  great  services  to  medicine  by  combining 
more  closely  than  had  ever  been  done,  the  technique  of  the 
chemist  with  the  in  vivo  experiments  of  the  biologist.  His 
guiding  principle,  which  dominates  his  whole  career  and  is  to 
be  found  in  his  earliest  works,  is  the  attempt  to  explain  vital 
actions  by  a  similar  mechanism  as  in  the  reactions  of  organic 
chemistry.  He  has  represented  by  stereochemical  formulae  the 
reciprocal  reactions  of  bodies  of  the  chemical  composition  of 
which  we  are  almost  entirely  ignorant,  the  toxins  and  antitoxins. 


288  MICROBES   AND  TOXINS 

He  has  disarticulated  protoplasm  into  atomic  groups,  both 
structural  and  functional,  which  act  like  the  organs  of  the 
cells;  he  has  extended  the  anatomy  and  physiology  of  the 
tissues  and  organs  by  imagining  a  sort  of  molecular  anatomy 
and  physiology  for  the  cell. 

The  cell  is  a  microcosm  in  which  multiple  functions  take 
place  by  continual  exchanges  between  the  protoplasm  and  the 
external  world.  Each  of  these  functions  is  represented  by  a 
group  or  "  side-chain,"  capable  of  entering  into  (chemical)  com- 
binations wijh  corresponding  groups  in  foreign  substances, 
food-stuffs,  poisons,  and  also  drugs.  All  the  vital  phenomena 
can  be  reduced  to  a  series  of  such  exchanges,  which  are  nutri- 
tion phenomena  and  retain  this  character,  whether  it  is  a 
protein  which  is  being  assimilated,  or  a  serum  producing 
immunity,  or  quinine  killing  the  malaria  parasite. 

For  a  substance  to  act  on  a  cell,  whether  an  organ  cell  or  a 
microbe,  it  must  fix  itself  on  the  protoplasm.  Did  not  a 
philosopher  in  the  middle  ages  declare  that  drugs  ought  to 
have  points  or  hooks  to  enable  them  to  seize  upon  the  organs  ? 
There  is  scarcely  any  phenomenon  which  haunts  the  mind  of 
the  biologist  more  than  this  fixation,  which,  according  to  some, 
is  a  physical  phenomenon,  one  of  molecular  adhesion,  according 
to  others  the  chemical  interplay  of  the  side-chains. 

The  antitoxin  injected  into  a  patient  has  affinities  only  for 
the  corresponding  toxin.  But  the  arsenic  which  we  inject  into 
an  individual  suffering  from  sleeping-sickness  possesses  affinities 
both  for  the  parasites  and  for  the  cells  of  certain  organs.  It  is 
a  double-edged  weapon.  We  must  suppress,  or  at  least  atten- 
uate as  much  as  possible,  the  dangerous  affinity  in  favour  of 
the  useful  one,  a  problem  in  chemical  substitution.  The  task 
is  chiefly  met  with  in  connection  with  protozoal  diseases,  but 
it  is  not  impossible  to  aim  in  a  similar  fashion  at  the  bacterial 
infections. 

We  already  know  examples  of  these,  we  may  call  them, 
elective  affinities.  Ehrlich  himself  showed  long  ago,  that 
methylene-blue  possesses  a  special  affinity  for  the  living  nerve- 
fibres  ;  there  has  even  been  derived  from  this  in  vivo  staining,  a 


CHEMICAL   REMEDIES  289 

valuable  method  for  anatomical  analysis  as  follows :  the 
animal  is  injected  with  methylene-blue  and  is  then  killed,  and 
one  finds  in  sections  the  nerve-fibres  as  fine  blue  lines.  By 
injecting  the  same  dye  into  a  frog,  the  nervous  system  of  the 
parasites  of  its  body-cavity  can  be  stained  in  the  most  delicate 
way.  There  are  in  cells,  granules  which  are  stained  electively 
by  neutral-red,  others  by  pyrrol-blue.  One  colour  is  taken  up 
by  the  nerve-cell,  another  by  the  fatty  material,  and  these 
stains  are  more  or  less  specific.  Drug  treatment  ought  to  be 
imagined  in  the  same  way.  To  cure  syphilis,  it  is  necessary  to 
find  a  chemical  compound  which  will  "stain"  electively  the 
parasite  of  Schaudinn  without  staining  the  cells. 

It  is  curious  that  we  find  here  again  this  analogy  of  staining, 
so  often  brought  forward  by  both  Ehrlich  and  Bordet,  but  with 
different  significations.  It  is  still  too  soon  to  ask  which  is  true, 
the  chemical  or  the  physical  theory. 

The  great  merit  of  a  theory  is  its  usefulness,  and  Ehrlich's 
theory  has  certainly  stimulated  much  work.  It  is  not  necessary 
to  retain  all  the  scaffolding  once  the  house  is  built. 

The  new  impulse  of  chemiotherapy  is  scarcely  six  years  old. 
Research  has  followed  different  tracks  and  attached  itself  to 
different  chemical  substances  :  hence  there  arose  several 
methods  in  which  various  remedies  were  soon  combined  and 
alternated. 

Here  is  a  list  of  the  principal  remedies  tried  against  the 
protozoal  diseases. 

1.  Trypan-red  and  the  benzidine  colours. 

2.  The  Triphenylmethane  series :   malachite-green,  brilliant- 
green,  crystal-  violet,  victoria-blue,  parafuchsin,  tryparosane.  .  .  . 

3.  Antimony,   tartar  emetic,  etc.      In  historical  and  logical 
order  this  series  was  studied   after   arsenic   and   atoxyl,    the 
chemical  relationship  of  antimony  and  arsenic  suggesting  this 
trial.     Tartar  emetic  acts  on  the  parasite  of  sleeping  sickness  : 
it  has  been  employed  in  man.     Salmon  has  observed  that  it 
acts — slightly — in  syphilis. 

4.  The  arsenical  bodies  and  atoxyl.     Arsenic  is  one  of  the 
most  ancient  remedies.     Even  before  Dioscorides  and  Pliny, 

u 


290  MICROBES   AND  TOXINS 

the  Chinese  had  employed  it.  As  soon  as  it  was  found  that 
sleeping  sickness  had  as  its  cause  a  trypanosome,  the  arsenical 
treatment  was  applied  to  it.  But  arsenious  acid  was  too  toxic 
to  be  easily  employed,  and  the  discovery  of  trypan-red  and  the 
first  successes  reported  with  the  chromotherapy  might  have  led 
to  its  abandonment,  had  not  W.  Thomas  brought  again  into 
prominence  the  substance  atoxyl  in.  1905.  Atoxyl,  discovered 
by  Bechamp  in  1863,  hardly  entered  the  sphere  of  human 
medicine  until  1902,  when  it  was  employed  in  dermatology, 
both  by  intravenous  and  subcutaneous  inoculation.  Thomas 
has  the  credit  of  employing  it  against  sleeping  sickness. 

It  is  not,  as  was  first  thought,  metarsenical  aniline,  which 
contains  377  per  cent,  of  arsenic.  According  to  Ehrlich  and 
Bertheim,  it  is  the  monosodic  salt  of  paraminophenylarsenic 
acid,  but  it  is  still  the  compound  discovered  by  Bechamp. 

It  contains  24  per  cent,  of  arsenic.  It  is  a  white,  crystalline 
powder  soluble  in  water  and  easily  sterilized.  It  may  also  be 
applied  as  an  ointment.  Although  almost  thirty  times  less 
toxic  than  arsenious  acid  excessive  doses  produce  nephritic 
symptoms  and  above  all  disturbance  of  vision  reaching  even  to 
complete  blindness. 

Atoxyl  has  been  employed  in  the  trypanosome  infections 
and  in  several  spirillum  diseases,  such  as  recurrent  fever  and 
syphilis  :  it  exerts  both  a  prophylactic  and  a  curative  action. 
It  has  been  said  that  it  no  longer  acts  in  the  trypanosomes  in 
sleeping  sickness  once  these  have  penetrated  into  the  cerebro- 
spinal  fluid  :  but  according  to  several  observers  the  meninges 
are  in  general  quite  permeable  (L.  Martin).  In  vitro  it  acts 
neither  on  trypanosomes  nor  on  spirilla,  so  that  the  body  itself 
must  play  an  active  part  in  the  treatment. 

Atoxyl  would  be  a  perfect  remedy  were  it  non-toxic.  By 
ringing  the  changes  on  the  chemistry  of  this  arsenical  subject 
a  series  of  compounds  have  been  discovered  of  which  the  last 
is  1,500  times  less  toxic  than  the  first. 

Three  compounds  superior  to  atoxyl  have  been  discovered 
in  this  series  :  in  the  first  place  arsacetin,  which  is  four  times 
less  toxic  than  atoxyl  for  the  mouse  and  distinctly  less  toxic 


CHEMICAL   REMEDIES  291 

for  the  monkey;  secondly,  arsenophenylglycin  (arsenophenyl 
glycocollate  of  sodium),  two  to  four  times  less  toxic  than 
atoxyl  and  more  active  in  killing  the  parasites  and  in  pro- 
phylactic power.  Quite  recently  Strong  and  Teague  in  the 
Philippines  have  been  treating  surra  in  horses  with  arseno- 
phenylglycin on  the  scale  of  veterinary  practice  and  not  of 
laboratory  experiment,  and  have  cured  more  than  one-third  of 
the  animals,  besides  checking  the  epidemic.  In  the  series  of 
bodies  studied  by  Ehrlich  arsenophenylglycin  had  the  number 
418.  Finally  we  reach  the  body  numbered  606,  the  dioxy- 
diamidoarsenobenzol  (C12H12O2N2As2),  of  which  the  hydro- 
chloride  is  employed  as  the  celebrated  remedy  termed  "  606  " 
or  Salvarsan. 

5.  The  toxicity  of  these  chemical  remedies,  the  necessity  of 
treating  relapses  and  the  fear  of  rendering  habituated  to  the 
drug  both  the  patient  and  the  parasite,  have  suggested  the 
method  of  combined  or  more  correctly  alternated  medication 
(Laveran).  The  drugs  associated  may  be  of  the  same  series 
or  of  a  different  chemical  series  from  the  principal  drug.  In 
spite  of  the  good  qualities  of  atoxyl  arsenious  acid  still  retains 
the  first  place  in  the  alternated  treatment. 

Treatment  of  Sleeping-Sickness. — From  1906  on- 
wards atoxyl  was  the  remedy  most  employed  in  the  treatment 
of  sleeping-sickness.  R.  Koch  during  his  sojourn  in  the  Sese 
Islands  on  the  Victoria  Nyanza,  gave  half  a  gram  two  days  in 
succession  :  in  about  eight  hours  the  parasites  disappeared 
from  the  blood  and  from  the  swollen  glands.  They  remained 
absent  for  about  ten  days,  when  the  injections  were  repeated  • 
without  inconvenience  this  double  injection  was  continued 
every  six  days  for  from  four  to  six  months. 

Koch  observed  further  that  on  ceasing  the  injections  the 
parasites  reappeared  in  the  glands  after  a  minimum  period  of 
eleven  days ;  from  the  twenty-fifth  day  they  could  be  found  in 
about  2  5  per  cent,  of  the  patients  treated  as  above ;  they  then 
disappeared  and  after  the  sixtieth  day  were  not  to  be  found 
again.  But  the  disappearance  from  the  glands  was  not  a  final 
cure :  there  were  relapses.  The  prolonged  treatment,  how- 

u  2 


292  MICROBES   AND   TOXINS 

ever,  permitted  the  campaign  of  prophylaxis ;  the  patients 
thus  treated  had  no  longer  parasites  in  their  blood  and  could 
not  therefore  furnish  the  glossina  carriers  with  new  supplies  of 
virus. 

Mild  cases  were  cured  after  a  treatment  of  from  four  to  six 
months.  At  the  end  of  1907  Koch  calculated  that  the  death- 
rate  among  the  treated  individuals  was  between  a  tenth  and  a 
twentieth  of  what  it  had  been  among  the  non-treated. 

Relapses  occur  even  among  the  patients  treated  from  the 
earliest  period,  and  even  14  months  after  the  treatment  ceases. 
The  more  intense  the  treatment  the  later  the  relapse. 

Soamin  (atoxyl  with  one  molecule  of  water  in  addition)  was 
apparently  rather  less  toxic  than  atoxyl ;  arsenophenylglycine 
produces  pain  at  the  point  of  injection,  but  it  seems  to  act 
in  patients  in  whom  atoxyl  fails. 

Of  the  remedies  supplied  by  other  chemical  groups  the  best 
hitherto  is  tartar-emetic  injected  intravenously.  It  ought  to  be 
tried  when  atoxyl  fails  and  the  association  of  atoxyl  with  tartar- 
emetic  has  often  seemed  better  than  either  of  these  drugs 
alone.  With  one  injection  per  week  it  is  possible  to  keep  the 
blood  and  glands  free  from  trypanosomes.  The  combinations 
atoxyl  +  mercury  and  atoxyl  +  sulphide  of  mercury  (orpiment) 
have  been  tried ;  atoxyl  remains  the  basis  of  all  the  remedies. 

The  Treatment  of  Syphilis. — Experimental  study  of 
syphilis  is  quite  recent  and  dates  from  the  announcement  of 
Metchnikoff  and  Roux  that  the  disease  can  be  inoculated  with 
certainty  in  anthropoid  apes,  presenting  not  only  the  primary 
symptoms,  but  secondary  symptoms  as  in  man.  By  his 
discovery  of  the  specific  microbe,  Schaudinn  furnished  us 
(1905)  with  the  best  means  of  controlling  under  the  microscope 
both  inoculation  and  treatment.  For  centuries  the  idea  has 
prevailed  that  syphilis  is  exclusively  a  human  ailment ;  since 
1903  it  has  been  transferred  from  man  to  the  higher  apes  and 
from  these  to  the  lower,  and  now  it  has  been  inoculated  in 
rabbits  and  guinea-pigs.  Man  however  remains  the  chief. 

Six  years  ago  the  medical  world  would  have  been  astonished 
at  a  prediction  that  syphilis  and  the  trypanosome  diseases 


CHEMICAL   REMEDIES  293 

would  one  day  be  treated  practically  by  the  same  remedies. 
Yet  analogies  were  already  in  existence  to  act  as  a  guiding 
principle,  for  the  name  of  horse-syphilis  had  long  been  given 
to  a  trypanosome  disease,  dourine,  which  possesses  clinical 
resemblances.  It  was  Schaudinn,  again,  who  maintained 
from  his  study  of  the  facts  the  relationship  between  trypano- 
somes  and  spirochaetes ;  he  was  convinced  that  a  protozoon 
cannot  be  properly  known  until  its  biological  cycle  has  been 
followed  completely,  and  he  therefore  studied  the  haematozoa 
as  a  zoologist  and  found  in  the  development  of  the  same 
species  trypanosome  forms  and  spirochaetes.  When  the 
spirochaete  of  syphilis  came  under  his  eye,  he  was  not  only 
prepared  to  see  it  but  to  believe  it  and  maintain  it.  There 
was  great  discussion  in  the  scientific  world  as  to  the  relation- 
ships of  this  new  microbe  :  was  it  a  protozoon  or  a  bacterium  ? 
These  researches  had  the  happy  result  of  drawing  attention  to  a 
problem  which  concerns  medical  practice  much  more  closely 
than  was  guessed,  and  they  prepared  the  way  for  the  revolution 
in  the  treatment  of  syphilis. 

This  sprang  from  an  old  idea.  In  an  article  dated  1867  in 
the  Dictionnaire  des  Sciences  medicales^  one  may  read  that  arsenic 
occasionally  succeeds  in  completing  a  cure  after  mercury  and 
iodine  :  "  it  is  especially  the  symptoms  which  have  resisted  the 
former  treatment  which  yield  to  the  action  of  arsenic." 

The  arsenical  treatment  of  syphilis  is  an  adaptation  of  the 
arsenical  treatment  of  the  trypanosome  infections,  inspired  by 
the  ideas  of  Schaudinn  and  the  labours  of  Ehrlich. 

Salmon  was  the  first  to  undertake  in  Metchnikoffs  laboratory 
methodical  studies  with  atoxyl.  The  well-known  toxicity  of  this 
substance  confined  him  to  small  doses.  Arsenophenylglycine, 
employed  by  Alt  in  general  paralysis,  was  found  to  produce 
temporary  improvements.  It  was  in  December,  1909,  that 
Ehrlich  mentioned  for  the  first  time  a  new  substance  con- 
taining the  same  arsenobenzol  group  as  arsenophenylglycine, 
namely,  the  dioxydiamidoarsenobenzol,  famous  to-day  under 
the  name  of  "  606  "  or  Salvarsan. 

All  that  has  been  published  hitherto  on  Ehrlich's  remedy  con- 


294  MICROBES   AND   TOXINS 

firms  the  great  progress  it  has  realized.  It  is  to  be  hoped  that 
the  future  will  confirm  these  promises  and  make  of  this  drug 
one  of  the  great  victories  of  therapeutical  science. 

Observations  on  immunity  in  Protozoal  Diseases. 
Strains  resistant  to  Sera.  The  future  of  Chemio- 
therapy. — To  chemiotherapy,  also,  we  owe  much  new 
knowledge  on  the  immunity  to  the  protozoal  diseases,  and 
on  immunity  in  general. 

The  question  of  the  dose  is  here  of  capital  importance ;  the 
object  is  to  begin  and  to  finish  the  treatment  at  one  blow,  but 
it  must  be  a  sledge-hammer  stroke.  Small  doses  cure  for  a 
time  without  producing  immunity,  and  relapses  therefore  occur. 
In  the  treatment  of  a  relapse  the  trypanosome  is  found  to  be 
different.  Sometimes  it  becomes  more  sensitive  to  the  action 
of  the  drug.  Sometimes  it  gives  the  impression  "  that  as  the 
result  of  the  absorption  of  those  parasites  killed  by  the  atoxyl, 
the  body  acquires  a  degree  of  immunity  which  then  interferes 
with  their  normal  development "  (Ehrlich).  As  a  rule,  however, 
such  immunity  is  ephemeral. 

But  what  is  most  commonly  observed  in  following  the  course 
of  a  first  attack  on  to  a  relapse,  and  again  from  one  relapse  to 
another,  is  a  diminution  in  the  efficacy  of  the  remedy.  The 
question  is,  is  it  the  body  or  the  parasite  which  has  changed  ? 
We  find  it  is  the  parasite,  for  when  inoculated  now  into  a  fresh 
animal  it  still  resists  the  same  drug.  The  injection  of  a  drug 
in  small  doses  is  in  fact  the  best  way  to  render  a  trypanosome 
resistant  to  it,  and  further  to  create  a  strain  of  trypanosome  which 
resists  the  remedy  and  preserves  this  resistance  even  after 
hundreds  of  passages,  transmitting  it  as  an  acquired  character. 

Trypanosome  strains  have  been  created  in  the  laboratories 
resistant  to  trypanred,  to  benzidine  dyes,  atoxyl,  and  tartar- 
emetic.  The  resistance  is  a  group  resistance,  /.£.,  it  applies  to 
all  drugs  of  the  same  series  or  chemical  family  :  a  trypanosome 
resistant  to  trypanred  is  still  sensitive  to  the  arsenic  bodies ;  it 
resists,  however,  also  the  benzidine  dyes,  which  are  a  good  deal 
different  from  trypanred.  It  can  be  seen  by  testing  various 
drug-groups  against  the  same  resistant  strain  that  biology  agrees 


CHEMICAL  REMEDIES  295 

with  chemistry  in  putting  tartar-emetic,  i.e.,  antimony,  in  the 
same  group  as  arsenic. 

The  resistance  acquired  towards  the  bodies  of  one  group 
presents,  however,  different  degrees ;  thus,  in  the  arsenical 
group  a  strain  resistant  to  atoxyl  is  still  sensitive  to  arseno- 
phenylglycine ;  if  it  is  now  rendered  resistant  to  this  latter 
substance  it  still  remains  sensitive  to  tartar-emetic  and  arsenious 
acid.  Again  subjected  to  arsenious  acid  it  becomes  resistant 
to  tartar-emetic;  no  strain,  however,  has  yet  been  created 
resistant  to  arsenious  acid. 

These  observations  have  important  consequences  for  practice ; 
they  indicate  that  it  may  be  necessary  to  attack  the  same 
trypanosome  by  different  remedies  and  form  the  reason  for 
the  method  of  combined  or  alternated  drug  treatment.  The 
drugs  superpose  their  actions  on  the  parasite,  but  not  necessarily 
on  the  body,  because  their  toxicity  does  not  bear  entirely  on 
the  same  organ  cells.  Further,  the  different  degrees  of  resis- 
tance which  exist  towards  drugs  of  the  same  group  show  that 
it  is  proper  to  associate  atoxyl  and  arsenious  acid,  although 
they  are  both  arsenical  bodies.  Attacking  the  same  parasite 
with  several  remedies  is  doing  the  same,  says  Ehrlich,  as  the 
entomologist  who  pins  out  a  butterfly  with  several  pins. 

Although  the  difficulty  may  be  overcome  by  using  these 
combined  treatments  the  appearance  of  resistant  strains  is 
always  a  danger.  In  sleeping-sickness,  for  example,  there  is  only 
one  good  drug,  atoxyl,  to  destroy  the  trypanosome,  and  in 
presence  of  strains  resistant  to  this,  one  is  rather  helpless,  the 
other  remedies  being  either  too  weak  or  too  toxic.  The 
danger  would  be  aggravated  should  the  trypanosomes  preserve 
hereditarily  their  acquired  resistance  through  their  passages  in 
the  tsetse-fly  which  conveys  them.  It  would  represent  the 
creation  of  a  more  virulent  and  less  curable  disease,  not  only  in 
an  individual,  but  throughout  a  whole  country. 

Things  do  not  go  on  in  the  living  body  exactly  as  in  a 
test-tube.  This  fact,  so  often  referred  to  already,  must  be 
insisted  upon  again. 

There  are  certain  chemical  substances  which  act   on  the 


296  MICROBES   AND  TOXINS 

trypanosome  neither  in  vitro  nor  in  vivo ;  others  are  active  in 
both,  for  example,  tartar-emetic. 

Methylene-blue  kills  certain  spirochaetes  in  the  test-tube  in 
a  dilution  of  i  in  6,000,000,  but  has  no  action  in  the  body  of 
a  mouse  even  when  in  500  times  greater  concentration. 

Atoxyl  does,  not  act  in  vitro  but  acts  in  the  living  body. 
There  are  even  substances  which  in  the  body  stimulate  the 
multiplication  of  certain  parasites ;  one  must  here  reckon  with 
the  body  as  a  factor  as  well  as  the  parasite,  and  the  problem 
of  the  best  drug  to  use  both  in  the  first  attack  and  in  relapses 
is  a  new  one  for  each  species  of  trypanosomes,  and  for  each 
species  of  animal :  chemical  therapeutics  is  therefore  not 
likely  to  become  more  simple. 

When  an  animal  acquires  immunity  towards  a  bacterial 
infection,  there  develop,  as  we  have  seen,  antibodies^  as  manifested 
by  new  properties  of  the  serum,  bactericidal,  preventive,  and 
curative.  Is  it  the  same  in  protozoal  infections?  Do  the 
chemical  remedies  induce  the  formation  of  antibodies  ? 

The  serum  of  animals  infected  with  trypanosomes  possesses 
microbicide  and  preventive  properties,  weak  it  is  true,  but 
definite;  they  appear  in  the  course  of  chronic  or  subacute 
infections.  There  is  reason  to  believe  in  the  presence  of  an 
immune  body  analogous  to  those  known  in  antibacterial 
immunity ;  it  acts  by  inducing  a  phagocytosis  of  the  parasite. 

Strains  of  protozoa  may  arise  resistant  to  the  serum  of 
animals,  infected  or  provisionally  cured,  just  as  with  the 
chemical  remedies ;  in  many  cases  it  is  a  true  variety  forma- 
tion, for  the  acquired  character  can  be  transmitted  through 
several  generations  (counted  by  mouse  passage). 

The  antibodies  in  animals  infected  by  trypanosomes  originate 
doubtless  from  trypanosomes  destroyed  and  absorbed.  At  the 
same  time,  the  rapid  destruction  of  a  great  number  of  the 
parasites  may  flood  the  body  of  the  host  with  substances  which 
act  as  poisons — a  sort  of  endotoxins.  In  a  man  suffering  from 
sleeping  sickness,  a  disappearance  of  the  trypanosomes  under 
the  influence  of  atoxyl  is  often  followed  by  an  attack  of  fever 
(L.  Martin). 


CHEMICAL  REMEDIES  297 

A  dose  of  a  chemical  remedy,  itself  non-toxic,  sometimes 
kills  the  mice  while  destroying  their  parasites.  Ehrlich  warns 
against  treating  with  "  606 "  infants  suffering  from  the 
septicaemic  form  of  congenital  syphilis ;  he  fears  an  intoxica- 
tion with  substances  derived  from  the  suddenly  dissolved 
spirochsetes. 

The  paradox  that  a  patient  may  give  a  negative  Wassermann 
reaction  which  after  treatment  with  "606"  becomes  positive,  is  to 
be  explained,  according  to  Ehrlich,  by  the  production  of  anti- 
bodies as  a  consequence  of  a  non-curative  dose.  And  it  is  a 
local  production  of  endo toxin,  under  the  influence  of  arsenic 
or  mercury,  which  produces  the  sudden  revival  of  cutaneous 
eruptions  in  syphilis,  known  as  the  phenomenon  of  Herxheimer. 

These  observations  on  strains  resistant  to  drugs  and  to  sera 
have  thrown  light  on  the  question  of  the  virulence  of 
protozoa.  Bacteria  also  acquire  resistance  to  the  defensive 
powers  of  the  higher  animals ;  the  streptococci  and  the 
anthrax  bacilli  clothe  themselves  with  a  mucous  capsule ; 
the  typhoid  bacilli  become  insensitive  to  the  agglutinins  of 
the  patient  whom  they  are  infecting  j  other  bacteria  secrete 
agressins\  all  .these  are  manifestations  of  resistance.  This 
"  immunization  of  microbes  "  raises  the  thought  that  the  laws  of 
virulence  may  be  fundamentally  the  same  for  both  bacteria  and 
protozoa. 

There  is  nothing  to  exclude  the  possibility  of  a  drug-therapy 
in  bacterial  infections  also,  and  it  is  always  possible  that  our 
vaccines  of  to-day,  and  in  particular  our  sera,  marvellous 
discoveries  as  they  are,  and  originating  in  the  most  scientific 
empiricism,  may  give  place  to  physico-chemical  remedies  better 
defined  and  more  direct.  It  is  from  chemistry  that  we  have 
to  expect  advances  in  the  healing  art.  The  phrase  of  Duclaux 
is  very  apt  in  this  connection. 

"  With  Pasteur  chemistry  invaded  the  field  of  medicine 
probably  never  to  leave  it" 


GLOSSARY 


Actinpmycosis — an  infectious  disease  affecting  cattle  and  some- 
times man,  characterized  by  tumour  growths  of  the  jaw,  the 
lungs,  the  tongue,  &c.,  and  due  to  a  streptothrix^  q.v.  :  known 
popularly  as  woody-tongue. 

Algae— cellular  cryptogamaceous  plants  including  many  seaweeds. 

Amylase — a  ferment  capable  of  decomposing  starch. 

Ankylostoma — the  "  hook-worm,"  a  parasite  of  the  small  intestine, 
common  in  the  tropics  and  among  miners,  and  producing 
severe  anaemia,  the  "  miners'  anaemia." 

Annelids — a  class  of  Vermes,  or  worms,  including  the  common 
earthworm. 

Ascitic  fluid — the  fluid  producing  abdominal  dropsy. 

Ascus — an  enlarged  cell  of  a  fungus  in  which  the  spores  are 
developed,  usually  the  terminal  end  of  a  hypha  or  thread. 

Auto-intoxication — the  poisoning  of  the  body  by  materials  developed 
within  itse]f:  gout,  arterio-sclerosis,  &c.,  are  supposed  to  be 
auto-intoxications. 

Basidium— the  spore-bearing  hypha  (or  thread)  of  a  fungus. 

Bothriocephalus — a  broad  tape-worm  occasionally  infecting  man 
and  derived  from  the  ingestion  of  certain  fish  in  which  its 
cystic  stage  occurs. 

Botulismus — the  poisoning  produced  by  the  consumption  of  meat, 
particularly  in  the  form  of  sausages,  in  which  an  anaerobic 
bacillus,  the  B.  Botulinus  of  Van  Ermenghem,  has  grown  and 
produced  a  toxin,  the  botulismus  toxin. 

Brownian  movement — dancing  vibratile  movements  seen  among 
minute  particles,  even  of  inert  substances  such  as  charcoal, 
when  suspended  in  a  fluid  and  examined  under  the  microscope  : 
it  is  not  to  be  confused  with  true  motility. 

Calories — units  of  heat :  large  and  small  calories  distinguished  by 
capital  and  small  letters  ;  small  calory  is  the  amount  of  heat 
necessary  to  raise  I  gram  of  water  i°  centigrade  :  large  calory 
the  amount  necessary  to  raise  i  kilogram  i°  centigrade. 

299 


300  MICROBES   AND   TOXINS 

Carbohydrates — compounds  of  the  three  elements,  carbon,  oxygen, 
and  hydrogen  :  the  chief  representatives  are  the  sugars  and 
starches. 

Catalysts— substances  which  modify  the  rapidity  of  a  reaction 
without  forming  part  of  its  final  products  :  such  a  reaction  is 
termed  "catalysis"  (Berzelius,  1835) :  f°r  example  peroxide  of 
hydrogen,  H2O2,  is  decomposed  into  water,  and  oxygen  on 
contact  with  spongy  platinum,  or  platinum  in  a  state  of  fine 
division.  The  platinum  remains  unaltered  :  it  has  acted 
merely  by  its  presence. 

Chemiotaxis — a  sort  of  "  chemical  sense  "  by  means  of  which  living 
cells  seem  to  seek  the  substances  favourable  to  them,  and 
avoid  those  which  are  injurious.  Positive  and  negative 
chemiotaxis  are  indicated  by  movements  of  attraction  and 
repulsion  respectively,  *.<?.,  by  approach  or  flight. 

Chromatin — the  substance  composing  the  greater  part  of  the 
nucleus  of  cells,  so-called  because  it  stains  very  deeply  with 
certain  dyes. 

Chromidia — masses  or  networks  of  chromatin  distinct  from  the 
nucleus  itself. 

Colloids — Graham  gave  the  name  of  "  colloids  "  to  those  substances 
which  in  watery  solution  do  not  dialyse  (i.e.,  do  not  pass 
through  a  parchment  membrane  dipping  in  pure  water),  or 
dialyse  extremely  slowly  in  contrast  to  the  "  crystalloid " 
substances  which  rapidly  dialyse.  Types  of  these  two  classes 
are  gum  and  salt.  Nowadays  it  is  more  usual  to  speak  of 
"substances  in  the  colloidal  state"  than  of  " colloidal  sub- 
stances." The  "  colloidal  state  "  is  a  state  of  suspension  of  one 
substance  in  another  as  contrasted  with  true  solution.  For 
example  gutta-percha  forms  a  true  solution  in  alcohol,  a 
colloidal  solution  in  water.  In  the  animal  body  the  cells,  the 
cell-membranes,  and  the  body-fluids,  all  consist  of  colloidal 
solutions,  the  condition  and  activities  of  which  are  regulated 
by  physico-chemical  laws  :  the  colloids  are  thus  of  extreme 
interest  to  biologists. 

Conidium — a  spore  of  a  fungus  produced  asexually  and  borne  on  a 
special  branch. 

Cytoplasm — the  protoplasm  of  the  cell-body  as  distinguished  from 
the  nucleus,  the  protoplasm  of  which  is  known  as  nucleo- 
plasm. 

Daphnia — the  common  water-flea. 

Dialysis—  vide  Colloids. 

Diapedesis — a  phenomenon  discovered  by  Cohnheim  :  the  white 
corpuscles  of  the  blood  (leucocytes)  emigrate  from  the  interior 
of  the  blood-vessels  by  a  process  of  active  movement  not 
passive  expulsion.  They  insinuate  themselves  between  the 
cells  forming  the  walls  of  the  capillaries,  and  drag  themselves 
through  as  narrow  threads  to  recover  their  rounded  shape 
outside. 


GLOSSARY  301 

Diastases — also  known  as  ferments  or  enzymes :  they  are  sub- 
stances which  can  produce  fermentations  in  the  absence  of 
living  cells  :  their  chemical  nature  is  unknown  and  they  are 
defined  simply  by  their  activities :  they  occur  in  the  form  of 
organic  substances  of  indefinite  composition  which  are  soluble 
in  water.  In  the  embryo  of  grain  (barley,  &c.)  there  exists  a 
diastase  which  transforms  the  starch  of  the  grain  into  sugar  in 
the  presence  of  warmth  and  moisture,  the  process  of  "  malting." 
( Vide  Fermentation.) 

Electrolytes — bodies  which  in  solution  break  up  into  "ions" 
carrying  electrical  charges  which  are  equal  and  opposite.  For 
example,  common  salt  in  solution  in  water  breaks  up  into  a 
sodium  ion  carrying  a  charge  of  negative  electricity,  the  cation 
and  a  chlorine  ion  carrying  a  charge  of  positive  electricity  the 
anion.  The  presence  of  electrolytes  in  water  renders  it  a  good 
conductor  of  electricity,  hence  the  name. 

Enzymes — vide  Diastases. 

Epizootic — infectious  disease  occurring  widespread  among  animals 
— contrasted  with  "  epidemic "  in  which  the  disease  affects 
man. 

Fats — compounds  of  carbon,  oxygen,  and  hydrogen  in  the  form  of 
"  esters  "  of  glycerine,  i.e.,  compounds  of  glycerine  with  a  fatty 
acid.  Palmitin,  stearin,  and  olein  are  fats  in  which  glycerine 
is  combined  with  palmitic,  stearic,  and  oleic  acids.  The 
natural  fats  are  mixtures  in  varying  proportions  of  palmitin, 
stearin,  and  olein.  The  "  saponification "  of  fats  is  their 
decomposition  into  the  two  elements  glycerine  and  fatty  acid, 
and  the  combination  of  the  latter  with  sodium  or  potassium 
("  salting  out "  in  the  language  of  soap  manufacture).  Soap  is 
thus  a  salt  of  sodium  or  potassium  with  a  fatty  acid,  e.g., 
stearate  of  sodium.  By  saponification  100  grams  of  fat  can 
furnish  90  grams  of  fatty  acid. 

Fermentation — a  chemical  transformation  produced  by  the  action 
of  living  cells,  e.g.,  the  cells  of  the  yeast  of  beer,  or  by  the 
action  of  the  secretions  of  these  cells  (diastases),  e.g.,  zymase 
extracted  from  yeast  cells.  A  typical  fermentation  is  that  of 
sugar  by  yeast  in  which  the  sugar  is  broken  up  into  carbonic 
acid,  water,  and  alcohol. 

Fibrin — a  protein  (more  exactly,  a  globulin)  which  forms  the  clot  in 
coagulated  blood.  It  does  not  exist  in  circulating  blood  in  the 
living  animal,  but  is  derived  from  a  different  substance  which 
exists  in  this  and  is  termed  fibrinogen.  "  Defibrinated  "  blood 
is  blood  from  which  the  fibrin  is  removed  as  it  forms  by 
whipping  with  a  bunch  of  twigs  immediately  after  the  blood  is 
drawn.  Such  blood  is  no  longer  capable  of  coagulation. 

Flora — the  aggregate  of  plants  growing  without  cultivation  in  a 
given  district  or  indigenous  to  a  particular  geological  formation : 
hence  applied  to  the  aggregate  of  bacterial  species  inhabiting 
the  intestine,  the  mouth,  &c. 


302  MICROBES   AND  TOXINS 

Gametes — sexually  differentiated  cells  which  unite  to  form  the 
fertilized  cell,  the  zygote. 

Glucosides — substances  which  are  capable  of  decomposition  into  a 
sugar  (glucose),  and  various  other  organic  substances,  alcohol, 
phenol,  aldehyde,  &c.  For  example,  aniygdalin  the  active 
principle  of  bitter  almonds  breaks  up  into  glucose  benzaldehyde 
(the  odorous  constituent)  and  hydrocyanic  (prussic)  acid. 
Many  poisonous  plants  contain  glucosides  as  their  active 
principle. 

Glycogen — a  carbohydrate  analogous  to  the  starches,  stored  by  the 
body  in  the  cells  of  the  liver  and  muscles,  and  drawn  upon  by 
the  body  for  the  supply  of  sugar  consumed  by  the  cells  during 
muscular  work  and  other  activities. 

Humours — body-fluids,  for  example  the  aqueous  and  vitreous 
humours  of  the  eyeball. 

Incubation — the  time  elapsing  between  the  moment  of  penetration 
of  the  body  by  a  virus  (microbe  or  toxin)  and  the  moment  when 
the  first  symptoms  appear. 

Indol — Among  the  products  of  the  digestion  and  putrefaction  of 
proteins  (q.v.)  there  appears  a  substance,  tryptophane,  from 
which  the  indol  bodies  are  derived,  indol  and  skatol  (indicated 
by  various  colour  reactions  depending  upon  their  relationship 
to  indigo)  :  also  a  substance,  tyrosin,  from  which  are  derived 
paracresol  z.ndi  phenol  (carbolic  acid). 

Inflammation — the  reaction  of  living  tissues  to  injuries  and 
infections.  The  essential  fact  in  Inflammation  is  the  activity 
of  cells,  the  phagocytes  which  are  capable  of  taking  up 
substances  and  digesting  them. 

Isomeric — with  different  chemical  or  physical  properties  but 
containing  the  same  quantity  of  chemical  elements  in  the 
molecule  :  the  difference  is  due  to  the  different  arrangement  in 
space  of  the  atoms  in  the  molecule. 

Lipoids — a  group  of  substances  such  as  lecithin,  cerebrin,  protagon 
cholesterin,  possessing  certain  of  the  properties  of  fats  {\ITTOV — 
fat).  The  name  was  introduced  by  Overton  to  indicate  the 
possession  by  these  bodies  of  powers  of  solution  similar  to  fats, 
in  particular  for  anaesthetics.  According  to  Overton  the  outer- 
most layer  of  protoplasm  in  a  cell  consists  of  lipoid  substances 
and  the  cell  absorbs  only  those  materials  which  are  soluble  in 
the  lipoids. 

Medulla — the  medulla  oblongata  or  bulb  is  the  continuation  of  the 
spinal  cord  within  the  skull  before  its  junction  with  the  brain  : 
it  contains  the  nuclei  of  certain  cranial  nerves  regulating  the 
respiration  and  the  heart-beat. 

Molluscum  contagiosum — an  infectious  skin  disease  characterized 
by  small  teat-like  protuberances. 

Mucedineae — fungi  of  the  group  to  which  the  ordinary  white  mould 
mucor  mucedo  belongs. 


GLOSSARY  303 

Mutations — sudden   variations    appearing    in    individuals   of  one 

species  and  capable  of  transmission  to  their  descendants. 
Mycetozoa  or   Myxomycetes — protozoa    with    naked    protoplasm 

occurring  on  damp  surfaces  in  the  form  of  jelly-like  masses, 

living    on   organic   debris,  arncebiform  without  mycelium  but 

later  plant-like. 
Myeloplax — a    multinucleated   cell   occurring  in  the    marrow  of 

bones. 
Nematodes — a    class  of  worms  with  thread-like  body,  mouth  and 

intestinal  canal,  including  the  parasitic  thread-worms,  &c. 
Neuroglia— cells  in  the  nervous  system  which  correspond  to  the 

connective  tissue  cells  in  other  organs. 
Nitrification — the  transformation  of  salts  of  ammonia  (chiefly  the 

sulphate  and  the  carbonate)  into  nitrites  and  nitrates  under  the 

influence  of  the  nitrifying  bacteria. 
Plasma — the  fluid   portion   of  the  blood  freed  from  the  red  and 

white  corpuscles  which  float  in  it  :  it  contains  fibrinogen   but, 

not  having  been  allowed  to  coagulate,  no  fibrin  (vide  Fibrin). 
Proteins — substances  resembling  egg-white  and  containing  nitrogen, 

carbon,  oxygen,  hydrogen,  and  sulphur. 
Ptomaines — products   of    putrefaction,   some   of  them   poisonous, 

isolated  from  putrefying   organic   matter  (first   discovered  in 

dead  bodies). 
Putrefaction — the  decomposition  of  protein  substances  by  microbes 

and  their  ferments  with  the  production  of  gas,  foul  smells,  and 

sometimes  poisonous   substances  :  as  a  result  of  putrefaction 

organic  matter  is  restored  to  the  state  of  inorganic  elements. 
Septic— derived  from  o-€7r<m — corruption  and  applied  to  material 

which  can  produce  or  undergo  bacterial  infection,  e.g.,  a  septic 

wound,   a   septic    instrument,  a  septic   dressing.     Hence  the 

words  "  antiseptic,"  a  substance  acting  against  this  action,  and 

"aseptic,"  the  absence  of  sepsis.    In  modern  surgery  antisepsis 

has  given  place  to  asepsis. 
Serum — the  clear  fluid  expressed  from  the  clot  of  coagulated  blood  : 

it  represents  the  fluid  portion  of  the  blood  minus  the  cells  and 

minus  the  fibrin  which  forms  the  clot  (vide  Fibrin). 
Stereochemical— the    chemistry    of   matter    may    be    treated    as 

depending    on   arrangements   of  the  atom   in   space   in   the 

molecule  (vide  isomeric). 
Sucrase — sugar-splitting  ferment. 
Symbiosis — the    living    together    of    dissimilar    organisms    each 

dependent  on   the   other  :  the  best  examples  are  the  lichens 

where  a  fungus  and  an  algae  live  together. 
Urease— urea-splitting    ferment :    producing    ammonia    from   the 

urea,  the  main  nitrogenous  constituent  of  urine. 
Vagus— the  nerve  supplying  the  heart,  lungs,  and  stomach. 
Vitalism — the  modern  application  is  to  a  theory  which  postulates, 

at  least  provisionally,  some  other  activity  in  the  life  processes 


304,  MICROBES   AND  TOXINS 

than  mechanical,  physical,  and  chemical  forces.  The  theory 
of  immunity  as  being  due  to  phagocytosis  calls  in  the  activity 
of  living  cells  the  mechanism  of  which  is  not  known.  It  is 
certain,  however,  that  in  vital  activities  it  is  not  necessary  to 
assume  anything  beyond  physico-chemical  phenomena  which 
will  some  day  become  clear.  Biologists  no  longer  talk  of  a 
"vital  principle." 
Zymase — vide  Fermentation. 


INDEX 


INDEX 


ABRIN,  168 

Acidity,  inhibiting  power,  IO,  42 

Acidophilus,  35 

Acinetians,  1 21 

Actinians,  venom  of,  178,  234,  238 

Actino-congestin  :  see  Congestin. 

Adulteration,  detection  of,  255 

Aerobes,  80 

in  fermentation,  5 

in  putrefaction,  10 
Agglutination,     Bordet's     explana- 
tion, 221 

diagnosis  by,  252 
Agglutinin,  development  of,  193 

power  of,  253 
Agressins,  231 
Albuminous  materials,  putrefaction 

of,  9 

Alcohol  production,  82 
Alexine  :  see  Complement. 
Algae,  299 

bacteria  relationship,  63 

coals,  7 

nitrogen  fixation,  20 
Algal  coals  :  see  Bogheads. 
Amboceptor  :  see  Immune-body. 
Amoebae,  culture,  105 

digestion,  99 

excretion,  101 

infection  among,  121 

inflammation  in,  134 

pathogenic,  142 
Amylase,  299 
Anaerobes,  80 

in  fermentation,  5 

in  putrefaction,  IO 
Anaphylaxis,  233 
Ankylostoma,  299 
Annelids,  299 
"  Antagonistic  force,"  IO 


Anthracosis,  47,  127 
Anthrax,  immunity,  207 

infection,  122 

vaccination,  270 

bacillus,    antagonistic    microbes, 
125 

encapsulation,  57 

involution  forms,  59 

light  effect  on,  96 

lysin,  87 

phagocytosis  of,  196 

respiration,  84 

virulence,  117 

virus  attenuation,  1 19 
Antianaphylaxis,  242 
Antibodies,  193,  248 

immunity,  207 
Antiendotoxins,  171 
Antigens,  193 

Bordet's  researches,  218 
Antinomycosis,  299 
Antisensibilism,  247 
Antitoxins,  168,  210,  227 
Apotoxine,  239 
Arachnolysin,  179 
Arsacetin,  290 
Arsenic  chemiotherapy,  289 
Arsenophenylglycin,  291 
Arrhenius  immunity  theory,  216 
Arthraspores,  59 
Arthus'  phenomenon,  234 
Ascarides,  respiration,  101 
Ascitic  fluid,  299 
Ascus,  229 
Aseptic  breeding,  30 
Aspergillus,  52 

Niger,  fermentation,  5 

nutrition,  73,  75 

respiration,  84 

zinc  absorption,  165 


3°7 


308 


INDEX 


Atoxyl,  290 
Auto-infection,  123,  88 
Auto-intoxication,  299 
Avian  plague,  154 
Axolotl,  infection,  134 
Azotobacter,  19 


BACILLUS,  55:  for  bacilli  of  different 

diseases  see  under  disease, 
acidi  paralactici,  42,  44 
acidophilus,  35 
amylobacter,  6,  7 
asterosporus,  64 
bifidus,  35 

chemical  properties,  44 

inhibiting  power,  42 
botulinus,  299,  37 

toxin,  159 
Blitschlii,  nucleus,  65 

reproduction,  68 

size,  150 
coli,  culture  of,  89 

in  intestine,  34,  36,  37 

mutation  of,  62 

para-,  115 

putrefaction  inhibited,  41 
cyanogenes,  pigment  of,  92 
exilis,  35 
lactis  serogenes,  34 

putrefaction  inhibited,  41 
maximus  buccalis,  65,  66 
megatherium,  109 
mesentericus,  35,  109 
perfringens,  inhibiting  action,  42 

in  intestine,  34,  36 
prodigiosus,  pigment,  92 

pleomorphism,  59 
putrificus,  Bienstock,  II 

inhibiting  action,  41 

in  intestine,  36 
pyocyaneus,     inhibiting     action, 

125 

Pfeiffer's  phenomenon,  204 

pigment,  93 
radicicola,  21 
ramosus,  58 
III.  of  Rodella,  34 
sporogenes,  36 
sporonema,  reproduction,  68 

cold  effect,  96 
subtilis,  involution  forms,  59 

respiration,  84 


Bacteria,  55 

alimentation,  77 

chemical  composition,  Jo 

diseases,  107 

heat  effect,  94 

light  effect,  96 

luminous,  89 

nitrogen  fixation,  19 

organic  cycles,  41 

pleomorphism,  59 

respiration,  80 

toxins,  1 68 
Bacteriaceae,  63 
Bacterial  beds,  17 
Bacteriolysis,  194 
Bacterio-purpurine,  85 
Bacterio-therapy,  273 

intestinal,  42 
Bacteriotropins, 
Bacterium  aceti,  5 

butyricum,  respiration,  80,  84,  85 

gammari,  64 

phosphoreum,  90 

sorbose,  79 

zopfii,  60 
Basidium,  299 
Beggiatoa,  63 

nutrition,  78 
Behring,  immunity  experiment,  201 

on  intestinal  permeability,  47 
Bellamy,  fermentation  experiment,  82 
Bengal  rose,  bactericidal  action,  97 
Bertrand,      G. ,      experiments      on 

medium,  79 

Besredka,  anaphylaxis  theory,  241, 
247 

endotoxins,  169 
Bienstock,  putrefaction  experiments 

10,  41 

Bier,  phagocytic  therapy,  284 
Bitumen,  bacterial  origin,  8 
Bleeding,  post,  92 
Blood,  pathogenic  microbes  in,  251 
Bloodstains,  detection  of,  255 
Blue  algae :  see  Cynnophyceae. 
Bogdanoff,  aseptic  breeding,  31 
Bogheads,  formation,  8 
Bordet,  Ch.,  agglutination,  254 
chemiotaxis,  136 
complement  fixation,  256 
haemolysis,  183 
immunity  theory,  218 
pleuropneumonia,  154 


INDEX 


309 


Bordet-Gengou  reaction,  220 
Borrel,  anti-tetanic  serum,  276 

pleuropneumonia,  154 
Bothriocephalus,  299 
Botulismus,  299 
Brownian  movement,  299 
Bruck,  an ti tuberculin  of,  177 
Buchner,   H.,   antitoxin  formation, 

210 

tubercle  infection,  125 
Bufotaline,  180 
Bufotenine,  180 
Bulgarian  bacillus,  44 
Butschlii,  on  bacterial  nucleus,  65 

CAGNIARD  DE  LA  TOUR,  cold  effect 

on  bacteria,  96 
Calcicolse,  22 
Calcifugse,  22 

Calmette,     antitoxin    experiments, 
212,  228 

ophthalmo-reaction,  262 
Carbohydrates,  300 

fermentation,  5 

synthesis,  3 

Cattle  plague  :  see  Rinderpest. 
Celluloses,  fermentation,  5,  6 
Centipedes,  venom,  179 
Cerebrin,  302 

Charrin,  immunity  experiments,  200 
Chemiotaxis,  300,  136 
Chemiotherapy,  285 
Cheyne,  Watson,    infection  experi- 
ments, 124 

Chlorella,  nitrogen  fixation,  20 
Chlorophyll,  3,  99 
Cholera,  anticholera  serum,  278 

infection,  122 

vaccination,  271 
vibrio,  40 

agglutination  test,  254 

endotoxin,  170 

flagella,  56 

microbes  favourable  to,  125 

phagocytosis,  202 

pseudo-,  114 

toxin,  171 

virulence  diminution,  117 
Cholesterin,  302 

lecithin  reaction,  185 

poisons  reaction,  225,  226 
Chonkewitch,      immunity     experi- 
ments, 208 


Chromatin,  300 

Chromidia,  65,  300 

Citron,  immunity  experiments,  208 

Cladothrix  dichotoma, 

Clostridium,  55 

pasteurianum,   nitrogen  fixation, 

19 

Coagulins,  248 

Coal,  bacterial  formation,  7 

Cobra  venom,  181 

Cocci,  55 

Coccidia,  life  cycle,  103 

parasitism,  104 

Cochin,  Denys,  fermentation,  83 
Cohen,  microbial   heat  production, 

89 

Cohendry,  aseptic  breeding,  32 
Cohn,  stability  of  species,  60 
Cohnheim,  inflammation,  131 
Colloids,  300 

immunity  analogies,  224 
Comma  bacillus,  40 
Complement,  183 

bactericidal  properties,  194 

Ehrlich's  theory,  215 

non-specificity,  256 

fixation,  256 

Complementophil  group,  215 
Congestion,  178,  234 

Richet's  researches,  238 
Conidium,  300,  52 
Conjunctis-reaction,  178 
Conradi :  inhibition,  88 
Contagious  diseases,  121 
Costia  necatrix,  103 
Cow-pox,  267 
Crenothrix  polyspora,  78 
Crepitin,  238 
Crotin,  168 
Cuti-reaction,  178 
Cyanophyceae,  bacteria  relationship, 

.63 

nitrogen  fixation,  20 
Cystococcus,  nitrogen  fixation,  20 
Cytodiagnosis,  252 
Cytophil  group,  215 
Cytoplasm,  300 


DANYSZ-DUNGERN  PHENOMENON, 

215,  222 
Daphnia,  300 

infection  by  monospora,  133 


310 


INDEX 


Denitrification,  12 
Diagnosis,  microbial,  250 
Diapedesis,  300 
Diarrhoea,  infantile,  38,  43 
Diastases,  301 
secretion,  86 
toxins  compared,  160 
Diplococci,  55 
Diphtheria,    antidiphtheric   serum, 

274 

bacillus,  lysin,  87 
pleomorphism,  59 
pseudo-,  113 
toxin,  158,  159 

virulence,  increase  and  diminu- 
tion of,  1 1 6,  117 

Dubois,  B.,  bacterial  luminosity,  91 
Duclaux,  fermentation,  83 
Dumas,  J.  B.,  fermentation,  82 
Dysentery,    antidysenteric     serum, 

279 

bacillus,  endotoxin,  170 
pseudo-,  114 
receptive  cell,  127 


EHRLICH,  chemiotherapy  research, 
287. 

immunity  theory,  21 1,  213 

serum  titration,  275 

side-chain  theory,  211 
Endospores,  59 
Endotoxins,  158,  168 
Engelmann ,  respiration  experiments, 

80,  85 

Entamoeba  histolytica,  145 
Enterococci,  35 

inhibiting  power,  42 
Enterokinase,  199 
Enzymes :  see  Diastases. 
Eosin,  bactericidal  action,  97 
Epidemics,  120 
Epitheliosis  virus,  128 
Erythrosin,  bactericidal  action,  97 
Evolution : 

bacterial,  108 

serum  action,  evidence  of,  256 

FACULTATIVE  BACTERIA,  80 
Falloise,  intestinal  putrefaction,  38 
Fats,  301 

fermentation,  5 

resistance  to  decomposition,  9 


Fermentation,  5,  301 

anaerobic  production,  82 
chemical  constitution  of  med- 
ium, 78,  79 
heat  production,  89 
phagocytosis  analogy,  195 

Ferments :  see  Diastases. 

Ferrau,  cholera  vaccination,  271 

Ferro-bacteria,  78 

Fibrin,  301 

Fibrinogen,  301 

Ficker,  intestinal  permeability,  48 

Filtration,  152 

Findel,  infection  experiments,  125 

Finkelstein,  on  diarrhoea,  38 

Fishes,  venoms  of,  178 

Foot-and-mouth  disease  virus,  155 

Fowl-cholera,  infection,  124 
bacillus,  lysin,  87 
size,  150 
virus  attenuation,  118 

Friedberger,    anaphylaxis     theory 
246 

Fungi,  bacteria  relationship,  62 
coals,  7 
sex,  66 

GALACTOSE,  fermentation,  79 

Germ-carriers,  124 

Giant  cell,  139 

Glossina  palpalis  :  see  Tsetse  fly 

Glucose,  fermentation,  78 

Glucosides,  302 

fermentation,  5,  6 
Glycogen,  302 
Granular     conjunctivitis,    see   Tra 

choma. 

Granulobacter,  7 
Gregarines,  104 

Gromia  oviformis,  digestion,  100 
Guarnieri  bodies,  155 

HAEMATOZOA,  pathogenic,  142 
Haemolysin,  179 

secretion  of,  86,  193 
Haemolysis,  Bordet's  theory,  221 

specific,  225 

of  venoms,  182 
Haffkine,  cholera  vaccination,  271 

plague  vaccination,  273 
Haptophorous  groups,  212 
Heat-production,  microbial,  89 
Heredity,  145 


INDEX 


311 


Herxheimer,  phenomenon  of,  297 
Hippuric  acid,  fermentation,  10 
Hoffmann's  bacillus,  113 
Hog-cholera,  46,  154 
Hogyes,  rabies  treatment,  269 
Horse-sickness,  154 
Humoral  immunity,  191 
Hymenoptera  venom,  179 

IMMUNE  BODY,  183,  195 

Ehrlich's  theory,  215 

specificity,  199 
Immunity,  187  et  seq. 

anaphylaxis,  240 
Incubation  of  disease,  128,  302 
Indol,  302 

toxicity,  37 
Infection,  121 

intestinal,  127 
Inflammation,  130,  302 
Infusoria,  51 

excretion.  101 

infection,  121 
Inoculation,  266 
Intermediate  substance :  see  Immune 

body. 

Intestine,  flora,  II,  27,  33 
Intra-dermo  reaction,  178 

JOCHMANN,  tuberculin  experiments, 
208 

KALA-AZAR,  144 

Khondiakon,  84 

Koch,  R.,  sleeping  sickness  treat- 
ment, 291 

specificity  of  disease,  112 
tubercle  phenomenon,  173 

Kurpjuweit,  on  inhibition,  88 

Kyes,  lecithids,  185 

LACCASE,  86 

Lactic  bacilli,  bacteriotherapy,  273 

fermentation,  43 

involution  forms,  59 
Laine,  artificial  nitre  beds,  1 6 
Lamblia  intestinalis,  104 
Lamprocystis,  78 
Lampyris  noctiluca,  91 
Latent  microbism,  123 
Lechartier,     fermentation     experi- 
ment, 82 
Lecithids,  184,  226 


Lecithin,  184,  226,  302 
Leguminosae  bacteria,  20  et  seq. 
Leishmania :  pathogenic,  142 

donovani,  varying  forms,  144 
Lentospora,  145 
Leprosy,  incubation  period,  128 

bacillus,  1 08 
Leptothrix  ochracea,  78 
Leucocidine,  87 
Leucogytes,  48,   136,   138  :  set  also 

Phagocytes. 

Levulose,  fermentation,  78 
Lignite,  bacterial  formation,  7 
Lipochrome,  92 
Light  production,  microbial,  89 
Lipoids,  1 66,  302 

colloids,  analogy  with,  224,  226 
Lithocolletis     caterpillars,     aseptic 

condition,  31 
Luciferine,  91 
Luciola  italica,  91 
Lumbricus,  inflammation  in,  133 
Lysius,  86,  248 

MACFAYDEN,  bacterial  extract,  169 

cold,  effect  on  microbes,  96 
Macrophages,  138,  197 
Madsen,  immunity  theory,  216 
Malaria,  infection,  123 

mosquito,  143 
parasite,  51,  104 

path  of  penetration,  126 

receptive  cell,  127 

reproduction,  66 
Mallein,  173 
Manure,  denitrification,  14 

fermentation,  7 
Marmorek's  serum,  280 
Martelly,  putrefaction  experiments, 

10 

Marx,  virus  fixe,  269 
Massart,  chemiotaxis,  136 
Massini,  bacterial  mutation,  62 
Maze,  fermentation  experiment,  82 
Meat,  adulteration  detection,  255 

putrefaction,  10 

Meningitis,   antimeningococcus    se- 
rum, 280 

bacteriotherapy,  274 
Meningococcus,  27 

pseudo,  114 

Mercaptan,  poisoning,  37 
Metachromatic  bodies,  64 


312 


INDEX 


Metchnikoff,  aseptic  breeding,  32 

cholera  bacillus,  40 

non-bacterial  life,  30 

pbagocytic  immunity,  197,  198 
Micrococci,  55 
Micrococcus  parvulus,  150 
Microphage,  138,  197 
Microsphaera,    amoebae    infection, 

121 

Miguel,  inhibition,  88 
Milk,  fermentation,  5 

putrefaction,  II,  41 
Molisch,  microbial  luminosity,  91 

respiration,  85 
Molluscum  contagiosum,  302 

virus,  155 
Monospora    tricuspidata,    Daphnia 

infection,  133 
Morgenroth,  antitoxin  experiments, 

212 
Moro,  aseptic  breeding  experiments, 

32 
Moulds,  52 

heat  effect,  95 

nitrogen  fixation,  20 
Mouth,  microbes  in,  27 
Mucedinae,  302 

nutrition,  74 
Mucor,  52 
Mucous  membranes,   microbes   in, 

26,46 
Mtintz,  nitrification  experiments 

.13,  16 
Mycelium,  52 
Mycetozoa,  303 

inflammation,  131 
Mycoderma  vini,  5 
Myeloplax,  303 
Myriapods,  9 
Mytilocongestin,  238 
Myxobolus  pfeifferi,  104,  145 
Myxomycetes :  see  Mycetozoa. 

NAGELI,  pleomorphism,  60 

Negri  bodies,  155 

Nepticula,  aseptic  condition,  31 

Neufeld,  bacteriotropins,  264 

Nicolle,  M.,  antibodies  theory,   248 

Nictric  ferment,  13,  74 

Nitrification.  12,  303 

Nitrobacter,  13 

Nitro-bacterium :  see  Nitric  ferment. 

Nitrogen  fixation,  18 


Nitrosococcus,  13 

Nitrosamonas,  13 

Nitrous  ferment,  13 

Nitsch,  virus  fixe  experiments,  269 

Nobecourt,  intestinal  putrefaction, 
38 

Nocard,  anti-tetanic  serum,  277 

Nose,  microbes  in,  26 

Nucleus,  bacterial,  64 

Nutrition,  microbial,  72 

Nuttall,  aseptic  breeding  experi- 
ments, 31 

OCULO-REACTION  :  see  Conjunctiro- 
reaction. 

Oi'dium  lactis,  II 

Omeliansky,  cellulose  fermenta- 
tion, 6 

Opsonins,  204 

Ornithodorous  monbata,  147 

Ornithorhynchus  venom,  180 

PANCREATIC    JUICE,     venom    re- 
action, 182 
Para,  bacteria,  112 
Para,  cresol,  302 
Parasitism,  102 
Passages,  method  of,  117 
Pasteur,  anaerobic  life,  80 

fermentation  experiment,  82 

immunity,  188 

microbial  association,  125 

non- bacterial  life,  28 

nutrition,  78 

pathogenic  microbes,  108 

pleuro-pneumonia  vaccination, 
270 

putrefaction,  10 

rabies  treatment,  268 

virulence,  117 
Pasteuria  ramosa,  57 
Pathogenic  microbes,  107 
Peat,  bacterial  formation,  7 
Pebrine,  146 
Pectose,  fermentation,  7 
Penicillium,  52 

nitrogen  fixation,  2O 

nutrition,  78 
Peptolytic  microbes,  10 
Petroleum,  bacterial  origin,  9 
Pettersson,    immunity    experiment, 

206 
Pfeiffer's  phenomenon,  201 


INDEX 


313 


Phagocytes,  135,  138 
Phagocytosis,  130,  191  (figs.),   IOO 

chemical  stimulation,  284 

immunity,  187 

toxins,  227 
Phagolysis,  198 
Pharynx,  microbes  in,  26 
Phenol,  302,  37 
Phosphorescence  (animal),  90 
Photodynamic  substances,  97 
Phycochrome,  63 
Pigments,  microbial,  92 
Piroplasma,  bigeminum,  147 
Piroplasmosis,  142 

infection,  123,  147 
Pirquet,  von,  serum -sickness,  234 

tuberculin,  178,  262 
Plasmodia,  142 
Plague,  anti-plague  serum,  279 

vaccination  for,  272 
Plants,  microbes  in,  28 
Plasma,  303 
Pleomorphism,  59 
Pleuropneumonia,  microbe,  153 

vaccination  for,  270 
Pneumococcus,  germ  carriers,  124 

in  throat,  27 
Pollen  coals,  1 8 
Portier,    anaphylaxis    experiments, 

234 

Precipitation  diagnosis,  254 
Precipitin,  193,  255 
Preisr,    tubercle    infection    experi- 
ment, 125 
Prophylaxis,  240 
Protagon,  230,  302 
Proteins,  303 

putrefaction,  9 
Proteolytic  microbes,  IO 
Proteus,  intestinal,  12,  37 

pleomorphism,  59 

putrefaction,  10 
Protozoa,  50 

diseases  due  to,  141,  286 

physiology  of,  98 
Pseudo-bacteria,  1 12 
Pseudomonas  lucifera,  90 
Ptomaines,  303 
Purple  bacteria,  85 
Putrefaction,  3,  303 

intestinal,  37 
Pyocyanase,  87 
Pyocyanin,  93 


Pyocyanolysin,  87 
Pyorrhea  alveolaris,  bacteriotherapy, 
273 

RABIES,  incubation  period,  128 

vaccination,  268 
-virus,  path  of  penetration,  127 

receptive  cell,  127 

virulence  increase,  117,  118 
Raulin,  nutrition  experiments,  74 
Raulin's  medium,  74 
Receptive  cells,  127 
Receptors  (Ehrlich),  212 
Recurrent  fever,  147 
Reproduction,  microbial,  67    102 
Respiration,  80 
Rhipicephalus  annulatus,  147 
Rhizopods,  pathogenic,  142 
Rhizopus  nigricans,  1 1 
Richet,    anaphylaxis    experiments, 

234 

Ricin,  168 
Rinderpest,  154 

sero- vaccination,  282 
Rivet,  intestinal  putrefaction,  38 
Roger,       immunity      experiments, 

200 
Roux,  immunity  experiments,  228, 

276 

pseudo-bacteria,  113 
serum  anaphylaxis,  241 
toxins,  1 60 

SACCHABOLYTIC  microbes,  10 
Sacred  host  :  see  Bleeding  host 
Salamander  venom,  180 
Saltpetre,  bacterial  formation,  15 
Salvarsan,  291,  293 
Sarcinae,  34 

pigment  production,  93 
Sarcocystine,  145 
Sarcosparidia,  104 
Scarlatina,  155 
Schaudinn,  nucleus,  bacterial,  65 

reproduction,  68 

syphilis,  118,  292 
Schick,  serum-sickness,  234 
Schizomycetes,  62 
Schizosaccharomycetes,  67 
Schloesing,      nitrification      experi- 
ment, 13 

Schottelius  :  see  Aseptic  breeding. 
Scorpion  venom,  179 


314 


INDEX 


Sea-urchin  venom,  178 

Sensibilisatrice  :  see  Immune  body. 

Sensibilism,  247 

Sensibilisinogen,  247 

Septic  tank  (sewage),  16 

Sero-agglutination,  252 

Sero-diagnosis,  255 

Serotherapy,  274 

Serovaccination,  245 

Serum,  274,  303 

Sewage  purification,  16 

Sex,  microbial,  67 

Sheep-pox,      sero-vaccination     for, 
281 

Side-chain  theory,  21 1 

Skatol,  302 

Skin,  bacteria  in,  26 

Sleeping    sickness  :     atoxyl     treat- 
ment, 290 
infection,  123 
treatment,  291 
trypanosome  of:  see  Trypano- 

soma  gambiense. 
tsetse-fly,  143 

Sleeswyk,     agglutination      experi- 
ments, 254 

Small-pox,  vaccines,  266 
virus,  128,  155 

Smith,  Th.,  anaphylaxis  phenome- 
non, 235,  248 

Snake  venom,  180 

Soamin,  292 

Sorbose,  bacterium  of,  79 

Spallanzani :  heat  effect  on  moulds, 

95 

Spider  venom,  179 

Spirilla,  55 

Spirillosis,  infection,  123,  148 

virus,  118 
Spirochaetes,  55 

diseases  due  to,  142 
Spore  coals,  formation,  8 
Spores,  58,  102 

heat  resistance,  95 
Staphylococci,  55 

in  skin,  26 
Staphylococcus  aurens,  92 

pyogenes,  37 
Staphylolysin,  87 
Starch  fermentation,  5 
Stegomyia  fasciata,  149 
Sterigmatocystes,  20 
Stichococcus,  20 


Stomach,  microbes  in,  27 
Streptococci,  34,  55 

anti-streptococcic  serum,  280 

encapsulated,  57 

in  skin,  26 

virulence,  116,  117 
Streptocolysin,  87 
Streptothriceae,  63 
Stylonychia  pustulata,  IOI 
Sucrase,  303 

Sugar,  fermentation,  5,  78 
soil-enrichment,  20 
Sulpho-bacteria,  alimentation,  78 

respiration,  85 
Sulphuretted   hydrogen    poisoning, 

37 

Sulphurous   bacteria  :    see   Sulpho- 
bacteria. 
Surgery,    phagocytic     therapy    in, 

283 

Swine-erysipelas  :  vaccination,  270 
-bacillus,  127 

virulence  increase,  118 
Symbiosis,  303 

Syphilis,  chemio-therapeutical  treat- 
ment, 292 
hereditary,  146 
sero- diagnosis,  258 
transmission,  148 
spirochoete,  path  of  penetration, 
126 


Takaki,   toxicity  experiments,    166 

226 

Tannase  fermentation,  6 
Tarantula  venom,  179 
Tartar-emetic,  in  sleeping  sickness, 

292 

Tetanolysin,  87,  215 
Tetanospasmin,  215 
Tetanus,  anti-tetanic  serum,  276 

favouring  factors,  125 

incubation  period,  128 

infection,  122 

latent,  123 
bacillus,  flagella,  56 

path  of  penetration,  163 

respiration,  84 

toxin,  159 

virulence  increase,  117 
Tetrodon  venom,  180 
Thalassine,  178 


INDEX 


315 


Thermophilic  bacteria,  94 
Thierfelder,  aseptic  breeding,  31 
Thomas,  W. ,  atoxyl  use,  290 
Tissier,    purification    experiments, 

10 

Toad  venom,  180 
Toxins,  157,  210 

phagocytosis,  227 
Toxogenin,  239 

Toxophore  group  [Ehrlich],  213 
Trachoma  virus,  155 
Trypanosoma,  51 

culture,  105 

diseases  due  to,  142,  290 

parasitism,  104 

sexual  reproduction,  66 
gambiense,  51,  105 

toxin,  145 
Trytophane,  302 
Tsetse  fly,  143 

Tubercle  bacillus,    chemical    com- 
position, 71 

culture  media,  76,  94 

detection,  112 

light,  effect  of,  97 

in  lung,  27 

para-,  115 

path  of  penetration,  127 

phagocytosis,  138 

virulence  increase,  117 
Tuberculin,  173 

chemical  composition,  71 

diagnosis  by,  260 

immunity,  208 

light,  effect  on,  97 
Tuberculosis,     antitubercukms     se- 
rum, 280 

antituberculous  vaccination,  272 

cutaneous  reactions,  262 

heredity,  146 

infection,  47,  121,  125 

tuberculin  treatment,  174 

tuberculin  diagnosis,  260 
bacillus  :  see  Tubercle-bacillus. 
Typhoid  bacillus,  agglutination  test, 
252 

blood  infection,  251,  46 

in  decayed  food,  37 

endotoxin,  170 

flagella,  56 

infection,  122 

lysin,  87 

para-,  115 


Typhoid  bacillus,  Pfeiffer's  pheno- 
menon, 204 
-fever,  germ  carriers,  124 

vaccination,  271 
Typhus  fever  bacillus,  120 
Ty rosin,  302 

ULTRA-MICROSCOPE,  151 
Urea  fermentation,  9 
Urease,  303 
Urine  fermentation,  9 

toxicity,  38 
Urobacteria,  9 
Urtilago  car  bo,  95 


VACCINATION,  265 

anaphylaxis,  234,  242 

continuous,  243 

sero-,  281 
Vaccines,  265 

passage  of,  118 

sensitized,  282 
Vaccinia  virus,  155 
Vaillard,  immunity  experiments, 

228 

Variolisation,  see  Inoculation 
Vaughan,  anaphylaxis  theory,  246 
Venoms,  178 

anti-venom  sera,  278 
Vibrio,  55 
Vibriolysin,  87 
Vibrion  septique,  putrefaction,  10 

respiration,  84 

Villes,  -Georges,     leguminosae    ex- 
periments, 20 

Vincent,  pathogenic  microbes,  109 
Virulence,  116 
Virus,  attenuation  of,  118 

filtrable,  150 

fixe,  268 
Vitalism,  303 


WALBRUM,  immunity  theory,  216 
Wassermann,  antrtuberculin,  177 
toxin  experiments,  166,  226 
reaction,  258 
Weever  fish  venom,  179 
Weigert,  65 

Welch's     bacillus  :      see     Bacillus 
perfringens, 


316  INDEX 

Wheeler,  anaphylactic  theory,  246  Yeasts,  fermentation,  5 
Widal,  biological  diagnosis,  252  milk  putrefaction,  II 

Willanen,  toxin  experiment,  212  nutrition,  77,  73 

Wollmann,  aseptic  breeding,  31  reproduction,  67 

Wolff-  Eisner,     ophthalmo-reaction,  Yellow  fever,  149,  154 

262  Yersin,  toxins,  159 
Wright,  opsonins  experiment,  204 


,  57 

YEASTS,  54  Zischeria,  aseptic  condition  of,  31 

chemical  composition,  70  Zygosaccharomycetes,  67 

cold,  effect  on,  96  Zymase,  83,  169 


M  Selection  from  the 
Catalogue  of 

G.  P.  PUTNAM'S  SONS 


Complete   Catalogues  sent 
on  application 


Putnam's 
Science    Series 


I.— The  Study  of  Man.  By  Professor  A.  C.  HADDON,  M.A.,  D.Sc.. 
M.R.I. A.  Fully  illustrated.  8",  net  $2.00. 

''  A  timely  and  useful  volume.  .  .  .  The  author  wields  a  pleasing  pen  and  knows 
how  to  make  the  subject  attractive.  .  .  .  The  work  is  calculated  to  spread  among  its 
readers  an  attraction  to  the  science  of  anthropology.  The  author's  observations  art 
exceedingly  genuine  and  his  descriptions  are  vivid." — London  Athenceum. 

2.— The  Groundwork  of  Science.  A  Study  of  Epistemology.  By 
ST.  GEORGE  MIVART,  F.R.S.  8°,  net  $1.75. 

14  The  book  is  cleverly  written  and  is  one  of  the  best  works  of  its  kind  ever  put  before 
the  public.  It  will  be  interesting  to  all  readers,  and  especially  to  those  interested  in  the 
study  of  science." — New  Haven  Leader. 

3. — Rivers  of  North  America.  A  Reading  Lesson  for  Students  of  Geo- 
graphy and  Geology.  By  ISRAEL  C.  RUSSELL,  Professor  of  Geology, 
University  of  Michigan,  author  of  "  Lakes  of  North  America,"  "  Gla- 
ciers of  North  America,"  '•  Volcanoes  of  North  America,"  etc.  Fully 
illustrated.  8°,  net  $2.00. 

44  There  has  not  been  in  the  last  few  years  until  the  present  book  any  authoritative, 
broad  resume  on  the  subject,  modified  and  deepened  as  it  has  been  by  modern  research 
and  reflection,  which  is  couched  in  language  suitable  for  the  multitude.  .  .  .  The  text 
is  as  entertaining  as  it  is  instructive." — Boston  Transcript. 

4.— Earth  Sculpture ;  or,  The  Origin  of  Land-Forms.  By  JAMES 
GEIKIE,  LL.D.,  D.C.L.,  F.R.S.,  etc.,  Murchison  Professor  of  Geology 
and  Mineralogy  in  the  University  of  Edinburgh ;  author  of  "  The 
Great  Ice  Age,"  etc.  Fully  illustrated.  8°,  net  $2.00. 

41  This  volume  is  the  best  popular  and  yet  scientific  treatment  we  know  of  of  the  ori- 
gin and  development  of  land-forms,  and  we  immediately  adopted  it  as  the  best  available 
text-book  for  a  college  course  in  physiography.  .  .  .  The  book  is  full  of  life  and  vigor* 
and  shows  the  sympathetic  touch  of  a  man  deeply  in  love  with  nature." — Science. 

5,— Volcanoes.  By  T.  G.  BONNEY,  F.R.S.,  University  College,  London. 
Fully  illustrated.  8°,  net  $2.00. 

"It  is  not  only  a  fine  piece  of  work  from  a  scientific  point  of  view,  but  it  is  uncom- 
monly attractive  to  the  general  reader,  and  is  likely  to  have  a  larger  sale  than  most  books 
of  its  class." — Springfield  Republican. 

6. — Bacteria  :  Especially  as  they  are  related  to  the  economy  of  nature,  to 
industrial  processes,  and  to  the  public  health.  By  GEORGE  NEWMAN, 
M.D.,  F.R.S.  (Edin.),  D.P.H.  (Camb.),  etc.,  Demonstrator  of  Bac- 
teriology in  King's  College,  London.  With  24  micro-photographs  of 
actual  organisms  and  over  70  other  illustrations.  8°,  net  $2.00. 

44  Dr.  Newman's  discussions  of  bacteria  and  disease,  of  immunity,  of  antitoxins,  and 
of  methods  of  disinfection,  are  illuminating,  and  are  to  be  commended  to  all  seeicing  in- 
formation on  these  points.  Any  discussion  of  bacteria  will  seem  technical  to  the  uniniti- 
ated, but  all  such  will  find  in  this  book  popular  treatment  and  scientific  accuracy  happilj 
combined."—  The  Dial. 


Book  of  Whales.    By  F.  E.  BEDDARD,  M.A.,F.R.S.    Illustrated 

8°.  $2.00. 

45  Mr.  Beddard  has  done  well  to  devote  a  whole  volume  to  whales.  They  are  worthy 
of  the  biographer  who  has  now  well  grouped  and  described  these  creatures.  The  general 
leader  will  not  find  the  volume  too  technical,  nor  has  the  author  failed  in  his  attempt  to 
produce  a  book  that  shall  be  acceptable  to  the  zoologist  and  the  naturalist."— N.  Y.  Times. 

8. — Comparative  Physiology  of  the  Brain  and  Comparative  Psy- 
chology. With  special  reference  to  the  Invertebrates.  By  JACQUES 
LOEB,  M.D.,  Professor  of  Physiology  in  the  University  of  Chicago. 
Illustrated.  8°.  $1.75. 

t  **  No  student  of  this  most  interesting  phase  of  the  problems  of  life  can  afford  to  remain 
in  ignorance  of  the  wide  range  of  facts  and  the  suggestive  series  of  interpretations  which 
Professor  Loeb  has  brought  together  in  this  volume."— JOSEPH  JASTROW,  in  the  Chicago 
Dial. 

$.— The  Stars.  By  Professor  SIMON  NEWCOMB,  U.S.N.,  Nautical  Al- 
manac Office,  and  Johns  Hopkins  University.  8°.  Illustrated.  Net. 
$2.00.  (By  mail,  $2.00.) 

**The  work  is  a  thoroughly  scientific  treatise  on  stars.  The  name  of  the  author  i» 
Sufficient  guarantee  of  scholarly  and  accurate  work."— Scientific  American. 

IO.— The  Basis  of  Social  Relations.  A  Study  in  Ethnic  Psychology.  By 
DANIEL  G.  BRINTON,  A.M.,  M.D.,  LL.D.,  Sc.D.,  Late  Professor  of 
American  Archaeology  and  Linguistics  in  the  University  of  Pennsyl- 
vania ;  Author  of  "History  of  Primitive  Religions,"  "Races  and 
Peoples,"  "  The  American  Race,"  etc.  Edited  by  LIVINGSTON  FAR- 
RAND,  Columbia  University.  8°.  Net,  $1.50  (By  mail,  $i. 60.) 

"  Professor  Brinton  his  shown  in  this  volume  an  intimate  and  appreciative  knowledge 
of  all  the  important  anthropological  theories.  No  one  seems  to  have  been  better  acquainted 
with  the  very  great  body  of  facts  represented  by  these  sciences."—  A m.  Journal  of 
Sociology. 

XX. — Experiments  on  Animals.  By  STEPHEN  PAGET.  With  an  Intro- 
duction by  Lord  Lister.  Illustrated.  8°.  Net,  $2.00.  (By  mail,  $2.20.) 

44  To  a  large  class  of  readers  this  presentation  will  be  attractive,  since  it  gives  to  them 
in  a  nut-shell  the  meat  of  a  hundred  scientific  dissertations  in  current  periodical  literature. 
The  volume  has  the  authoritative  sanction  of  Lord  Lister." — Boston  Transcript. 

12.— -Infection  and  Immunity.  With  Special  Reference  to  the  Preventioa 
of  Infectious  Diseases.  By  GEORGE  M.  STERNBERG,  M.D.,  LL.D., 
Surgeon-General  U.  S.  Army  (Retired).  Illustrated.  8°.  Net,  $1.75. 
(By  mail,  $1.90.) 

**  A  distinct  public  service  by  an  eminent  authority.  This  admirable  little  work  shouW 
bf!  a  part  of  the  prescribed  reading  of  the  head  of  every  institution  in  which  children  o| 
youths  are  gathered.  Conspicuously  useful."— N.  Y.  Times. 

13.— Fatigue.  By  A.  Mosso,  Professor  of  Physiology  in  the  University 
of  Turin,  Translated  by  MARGARET  DRUMMOND,  M.A.,  and  W.  B. 
DRUMMOND,  M.B.,  C.M.,  F.R.C.P.E.,  extra  Physician,  Royal  Hospital 
for  Sick  Children,  Edinburgh;  Author  of  "The  Child,  His  Naf^ 
and  Nurture."  Illustrated.  8°.  Net,  $1.50. 

**  A  book  for  the  student  and  for  the  instructor,  full  of  interest,  also  for  the  intelligent 
general  reader.  The  subject  constitutes  one  of  the  most  fascinating;  chapters  ii»  ««  Ju* 
tory  of  medical  science  and  of  philosophical  reseurc.li  "—Yorkshire  Post. 


14.— Earthquakes.  In  the  Light  of  the  New  Seismology.  By  CLARENCE 
E.  DUTTON,  Major,  U.  S.  A.  Illustrated.  8°.  Net,  $2.00. 

"  The  book  summarizes  the  results  of  the  men  who  have  accomplished  the  great 
tilings  in  their  pursuit  of  seismological  knowledge.  It  is  abundantly  illustrated  and  i» 
ills  a  place  unique  in  the  literature  of  modern  science." — Chicago  Tribune, 

15.— The  Nature  of  Man.  Studies  in  Optimistic  Philosophy.  By  ELi» 
METCHNIKOFF,  Professor  at  the  Pasteur  Institute.  Translation  and 
introduction  by  P.  CHAMBERS  MITCHELL,  M.A.,  D.Sc.  Oxon.  Illus- 
trated. 8°.  Net,  $1.50. 

"A  book  to  be  set  side  by  side  with  Hu^^'s  Essays,  whose  spirit  it  carries  a  step 
further  on  th^  long  road  towards  its  goal." — Mail  and  Express. 

16.— The  Hygiene  of  Nerves  and  Mind  in  Health  and  Disease.    By 

AUGUST  FOREL,  M.D.,  formerly  Professor  of  Psychiatry  in  the  Uni- 
versity of  Zurich.     Authorized  Translation.     8°.     Net,   $2.00. 

A  comprehensive  and  concise  summary  of  the  results  of  science  in  its  cnosen  field. 
Iti  authorship  is  a  guarantee  that  the  statements  made  are  authoritative  as  far  as  the 
statement  of  an  individual  can  be  so  regarded. 

17. — The  Prolongation  of  Life.    Optimistic  Essays.     By  £LIE  METCH- 
NIKOFF,  Sub-Director  o*  the   Pasteur  Institute.     Author   of   "  Tb« 
Nature  of  Man,"  etc.     8°.     Illustrated.     Net,  $2.50.    Popular  Edition. 
With  an  introduction  by  Prof.  CHARLES  S.  MINOT.     Net,  $1.75. 

In  his  new  work  Professor  Metchnikoff  expounds  at  greater  length,  in  the  light  of 
additional  knowledge  gained  in  the  last  few  years,  his  mam  thesis  that  human  life  is  not 
only  unnaturally  short  but  unnaturally  burdened  with  physical  and  mental  disabilities. 
He  analyzes  'he  causes  of  these  disharmonies  and  explains  his  reasons  for  hoping  that 
they  may  be  counteracted  by  a  rational  hygiene. 

1 8.— The  Solar  System.  A  Study  of  Recent  Observations.  By  Prof. 
CHARLES  LANE  POOR,  Professor  of  Astronomy  in  Columbia  University. 
8°.  Illustrated.  Net,  $2.00. 

The  subject  is  presented  in  untechnical  language  and  without  the  use  of  mathemaucs. 
Professor  Poor  shows  by  what  steps  the  precise  Knowledge  of  to-day  has  been  reached  and 
explains  the  marvellous  results  of  modern  methods  and  modern  observations. 

19.— Climate— Considered  Especially  in  Relation  to  Man.  By  ROBERT 
DECOURCY  WARD,  Assistant  Professor  of  Climatology  in  Harvard 
University.  8°.  Illustrated.  Net,  $2.00. 

This  volume  is  intended  for  persons  who  have  not  had  special  training  in  the  tech- 
nicalities of  climatology.  Climate  covers  a  wholly  different  field  from  that  included  in 
the  meteorological  text-books.  It  handles  broad  questions  of  climate  in  a  way  which  has 
not  been  attempted  in  a  single  volume  The  needs  of  the  teacher  and  student  have  been 
kept  constantly  in  mind. 

30. — Heredity.  By  J.  ARTHUR  THOMSON,  M.A.,  Professor  of  Natural 
History  in  the  University  of  Aberdeen  ;  Author  of  "The  Science  of 
Life,"  etc.  8°.  Illustrated.  Net,  $3.50. 

The  aim  of  this  work  is  to  expound,  in  a  simple  manner,  the  facts  cf  heredity  and 
Inheritance  as  at  present  known,  the  general  conclusions  which  have  been  securely 
established,  and  ihe  more  important  theories  which  have  been  formulated. 

21.— Age,  Growth,  and  Death.  By  CHARLES  S.  MINOT,  James  Still' 
man  Professor  of  Comparative  Anatomy  in  Harvard  University, 
President  of  the  Boston  Society  of  Natural  History,  and  Author  of 
*'  Human  Embryology,"  *' A  Laboratory  Text-book  of  Embryology," 
etc.  8°.  Illustrated.  $2.50  net. 

This  volume  deals  with  some  of  the  fundamental  problems  of  biology,  and  present! 
.  leries  of  views  (the  results  of  nearly  thirty  years  of  study),  which  the  author  hal 
"  *h*  first  time  in  systematic  form. 


£2. — The  Interpretation  of  Nature.    By  C.  LLOYD  MORGAN,  LL.D., 

F.R.S.     Crown  8vo.     Net,  $1.25. 

Dr.  Morgan  seeks  to  prove  that  a  belief  in  purpose  as  the  causal  reality  of  which 
nature  is  an  expression  is  not  inconsistent  with  a  full  and  whole-hearted  acceptance  of 
the  explanations  of  naturalism. 

23.— Mosquito  Life.  The  Habits  and  Life  Cycles  of  the  Known  Mos- 
quitoes of  the  United  States  ;  Methods  for  their  Control  ;  and  Keys  for 
Easy  Identification  of  the  Species  in  their  Various  Stages.  An  account 
based  on  the  investigation  of  the  late  James  William  Dupree,  Surgeon- 
General  of  Louisiana,  and  upon  the  original  observations  by  the  Writer. 
By  EVELYN  GROESBEECK  MITCHELL,  A.B.,  M.S.  With  64  Illustra- 
tions. Crown  8vo.  Net,  $2.00. 

This  volume  has  been  designed  to  meet  the  demand  of  the  constantly  increasing 
number  of  students  for  a  work  presenting  in  compact  form  the  essential  facts  so  far  made 
known  by  scientific  investigation  in  regard  to  the  different  phases  of  this,  as  is  now  con- 
ceded, important  and  highly  interesting  subject.  While  aiming  to  keep  within  reason- 
able bounds,  that  it  may  be  used  for  work  in  the  field  and  in  the  laboratory,  no  portion 
of  the  work  has  been  slighted,  or  fundamental  information  omitted,  in  the  endeavor  to 
carry  this  plan  into  effect. 

24. — Thinking,  Feeling,  Doing.  An  Introduction  to  Mental  Science. 
By  E.  W.  SCRIPTURE,  Ph.D.,  M.D.,  Assistant  Neurologist  Columbia 
University,  formerly  Director  of  the  Psychological  Laboratory  at  Yale 
University.  189  Illustrations.  2d  Edition,  Revised  and  Enlarged. 
Crown  8vo.  Net,  $1.75. 

u  The  chapters  on  Time  ai  i  Action,  Reaction  Time,  Thinking  Time,  Rhythmic 
Action,  and  Power  and  Will  are  most  interesting.  This  book  should  be  carefully  read 
by  every  one  who  desires  to  be  familiar  with  the  advances  made  in  the  study  of  the 
mind,  which  advances,  in  the  last  twenty-five  years,  have  been  quite  as  striking  and 
epoch-making  as  the  strides  made  in  the  more  material  lines  of  knowledge." — Jour. 
Amer.  Med.  Ass^n.,  Feb.  22,  1908.  , 

25. — The  World's  Gold.     By  L.  DE  LAUNAY,    Professor  at  the  Ecole 
Superieure    des   Mines.     Translated   by   Orlando   Cyprian  Williams. 
With  an  Introduction  by  Charles  A.  Conant,  author  of  "  History  of 
Modern  Banks  of  Issue,"  etc.     Crown  8vo.     Net,  $1.75. 
M.  de  Launay  is  a  professor  of  considerable  repute  not  only  in  France,  but  among 
scientists  throughout  the  world.     In  this  work  he  traces  the  various  uses  and  phases 
of  gold  ;  first,  its  geology  •  secondly,  its  extraction  ;  thirdly,  its  economic  value. 

26. — The  Interpretation  of  Radium.  By  FREDERICK  SODDY,  Lecturer 
in  Physical  Chemistry  in  the  University  of  Glasgow.  Crown  8vo. 
With  Diagrams.  Net,  $1.75. 

As  the  application  of  the  present  day  interpretation  of  Radium  (that  it  is  an  element 
undergoing  spontaneous  disintegration)  is  not  confined  to  the  physical  sciences,  but  has 
a  wide  and  general  bearing  upon  our  whole  outlook  on  Nature,  Mr.  Soddy  has  presented 
the  subject  in  non-technical  language,  so  that  the  ideas  involved  are  within  reach  of  the 
lay  reader.  No  effort  has  been  spared  to  get  to  the  root  of  the  matter  and  to  secure 
accuracy,  so  that  the  book  should  prove  serviceable  to  other  fields  of  science  and  investi- 
gation, as  well  as  to  the  general  public. 

27. — Criminal  Man.  According  to  the  Classification  of  CESARE  LOM- 
BROSO.  Briefly  Summarized  by  his  Daughter,  Gina  Lombroso  Ferrero. 
With  36  Illustrations  and  a  Bibliography  of  Lombroso's  Publications 
on  the  Subject.  Crown  8vo.  Net,  $2.00. 

Signora  Guglielmo  Ferrero's  resume  of  her  father's  work  on  criminal  anthropology  is 
specially  dedicated  to  all  those  whose  office  it  is  to  correct,  reform,  and  punish  the  crimi- 
nal, with  a  view  to  diminishing  the  injury  caused  to  society  by  his  anti-social  acts  ;  also 
to  superintendents,  teachers,  and  those  engaged  in  rescuing  orphans  and  children  of 
vicious  habits,  as  a  guide  in  checking  the  development  of  evil  germs  and  eliminating 
Inco'-n'spble  subjects  whose  example  is  a  source  of  corruption  to  other* 


28. — The  Origin  of  Life.  Being  an  Account  of  Experiments  with  Certaiu 
Superheated  Saline  Solutions  in  Hermetically  Sealed  Vessels.  By  H. 
CHARLTON  BASTIAN,  M.D.,  F.R.S.,  Emeritus  Professor  of  the  Princi- 
ples and  Practice  of  Medicine,  University  College,  London  ;  Author  of 
'*  The  Nature  and  Origin  of  Living  Matter,"  "  The  Evolution  of  Life," 
etc.  8vo.  With  10  Plates  Containing  6l  Illustrations  from  Photo- 
micrographs. $1.50  net. 

u  This  most  noteworthy  and  compelling  book.  .  .  .  The  question — both  as  to 
che  supposed  origin  of  life  once  and  for  all,  and  also  as  to  the  supposed  impassable  gap  of 
to-day — is  surpassed  in  interest  by  nothing  in  the  whole  range  of  physical  sciences  ;  if, 
indeed,  there  be  any  to  equal  it  whether  in  interest  or  in  moment  for  our  philosophy." 

The  Morning  Post. 

29. — The  Bacillus  of  Long  Life.  A  Manual  of  the  Preparation  and 
Souring  of  Milk  for  Dietary  Purposes ;  Together  with  an  Historical 
Account  of  the  Use  of  Fermented  Milks  from  the  Earliest  Times  to  the 
Present  Day,  and  their  Wonderful  Effect  in  the  Prolonging  of  Human 
Existence.  By  LOUDON  M.  DOUGLAS,  F.R.S.E.  8vo.  With  56 
Illustrations.  $1.50  net. 

This  book  has  been  designed  with  a  view  to  meet  an  extensive  demand  for  definite 
data  on  the  subject  of  Soured  Milks.  The  author  has  had  this  matter  brought  before 
him,  times  without  number,  by  those  inquiring  for  authentic  information  on  the  subject, 
and  he  has  therefore  considered  it  desirable  to  gather  together  such  information  as  is 
available  in  connection  with  ancient  and  modern  practice.  He  has  endeavored  to  pre- 
sent this  to  the  reader  in  concise  form. 

30. — The  Social  Evil.  With  Special  Reference  to  Conditions  Existing 
in  the  City  of  New  York.  A  Report  Prepared  in  1902  under  the  Di- 
rection of  the  Committee  of  Fifteen.  Second  Edition  Revised  with 
New  Material  Covering  the  Years  1902-191 1.  Edited  by  EDWIN  R.  A. 
SELJGMAN,  LL.D.,  McVickar  Professor  of  Political  Economy  in 
Columbia  University.  8vo.  $f.?5  net.  (By  mail^  $1.95.) 

A  study  that  is  far  from  being  of  merely  local  interest  and  application.  The  prob- 
lem is  considered  in  all  its  aspects  and,  for  this  purpose,  reference  has  been  made  to 
conditions  prevailing  in  other  communities  and  to  the  different  attempts  foreign  cities 
have  made  to  regulate  vice. 

31. — Microbes  and  Toxins.  By  ETIENNE  BURNET,  of  the  Pasteur  In- 
stitute, Paris.  With  an  Introduction  by  Elie  Metchnikoff,  Sub- 
Director  of  the  Pasteur  Institute,  Paris.  With  about  71  Illustrations. 

In  preparation  * 

The  Invisible  Spectrum.     By  Professor  C.  E.  MENDENHALL,  University 

of  Wisconsin. 
The  Physiology  and  Hygiene  of  Exercise.    By  Dr.  G.  L.  MEYLAN, 

Columbia  University. 

Other  volumes  to  be  announced  later 


UNIVERSITY  OF  CALIFORNIA 

MEDICAL  CENTER   LIBRARY 


THIS  BOOK  IS  DUE  ON  THE  LAST  DATE 
STAMPED  BELOW 

Books  not  returned  on  time  are  subject  to  a  fine  of  50c 
per  volume  after  the  third  day  overdue,  increasing  to 
$1.00  per  volume  after  the  sixth  day.  Books  not  in  de- 
mand may  be  renewed  if  application  is  made  before  expi- 
ration of  loan  period. 


QR41  Burn( 
B96b   Microbes 
1912   Dr. 


•?  r     o 

...  •  S  <: 


t,    E. 

and  to 
Charles 


ott. 


D7121 

ins.  Tr.   by 
Brodiuet   and    W. 


